WORLD INTELLECTUAL PROPERTY ORGANIZATION
International Bureau
per
INTERNATIONAL APPLICATION PUBLISHED UNDER THE PATENT COOPERATION TREATY (PCT)
(51) International Patent Classification 5 :
(11) International Publication Number:
WO 94/02136
A61K 31/395
Al
(43) International Publication Date:
3 February 1994 (03.02.94)
(21) International Application Number: PCT/US93/06678
(22) International Filing Date: 16 July 1993 (16.07.93)
(30) Priority data:
07/915,146
17 July 1992(17.07.92)
US
(71) Applicant (for all designated States except US): SMITH-
KLINE BEECHAM CORPORATION [US/US]; Cor-
porate Intellectual Property, UW2220, 709 Swedeland
Road, P.O. Box 1539, King of Prussia, PA 19406-0939
(US).
(72) Inventor; and
(75) Inventor/ Applicant (for US only) : LUENGO, Juan, Ignacio
[ES/US]; 701 Pondview Drive, Audubon, PA 19403
(US).
(74) Agents: STERCHO, Yuriy, P. et al.; SmithKline Beecham
Corporation, Corporate Intellectual Property, UW2220,
709 Swedeland Road, P.O. Box 1539, King of Prussia,
PA 19406-0939 (US).
(81) Designated States: AU, BB, BG, BR, BY, CA, CZ, FI, HU,
JP, KP, KR, KZ, LK, MG, MN, MW, NO, NZ, PL, RO,
RU, SD, SK, UA, US, VN, European patent (AT, BE,
CH, DE, DK, ES, FR, GB, GR, IE, IT, LU, MC, NL,
PT, SE), OAPI patent (BF, BJ, CF, CG, CI, CM, GA,
GN, ML, MR, NE, SN, TD, TG).
Published
With international search report.
(54) Title: RAPAMYCIN DERIVATIVES
(57) Abstract
Rapamycin derivatives; pharmaceutical compositions comprising such rapamycin derivatives and pharmaceutically ac-
ceptable carriers or diluents ; and methods of using such derivatives to inhibit pathogenic fungi growth, inhibit immunosuppres-
sion or treat carcinogenic tumors are disclosed.
FOR THE PURPOSES OF INFORMATION ONLY
Codes used to identify States party to the PCT on the front pages of pamphlets publishing international
applications under the PCT.
AT
Austria
FR
France
MR
Mauritania
AU
Australia
GA
Gabon
MW
Malawi
BB
Barbados
GB
United Kingdom
NE
Niger
BE
Belgium
GN
Guinea
NL
Netherlands
BF
Burkina Faso
GR
Greece
NO
Norway
BG
Bulgaria
HU
Hungary
NZ
New Zealand
Bj
Benin
IE
Ireland
PL
Poland
BR
Brazil
IT
Italy
PT
Portugal
BY
Belarus
JP
Japan
RO
Romania
CA
Canada
KP
Democratic People's Republic
RU
Russian Federation
CF
Central African Republic
of Korea
SO
Sudan
CG
Congo
KR
Republic of Korea
SE
Sweden
CH
Switzerland
KZ
Kazakhstan
SI
Slovenia
CI
Cote d'lvoire
LI
Liechtenstein
SK
Slovak Republic
CM
Cameroon
LK
Sri Lanka
SN
Senegal
CN
China
LU
Luxembourg
TD
Chad
cs
Czechoslovakia
LV
Latvia
TC
Togo
cz
Czech Republic
MC
Monaco
UA
Ukraine
DE
Germany
MG
Madagascar
US
United Slates of America
DK
Denmark
ML
Mali
UZ
Uzbekistan
ES
Spain
MN
Mongolia
VN
Viet Nam
FI
Finland
WO 94/02136
PCT/US93/06678
TITLE
RAPAMYCIN DERIVATIVES
BACKGROUND OF THE INVENTION
This invention relates to rapamycin derivatives,
pharmaceutical compositions comprising such derivatives,
and methods of treatment of pathogenic fungi, methods of
inducing immunosuppression and methods of treating
carcinogenic tumors utilizing such rapamycin
derivatives .
Rapamycin is a naturally occurring macrocyclic
triene antibiotic which can be produced by culturing an
organism in an aqueous nutrient medium. Its structure
may be illustrated as follows:
Figure I
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10
15
20
25
At least one rapamycin-producing strain of
Streptomyces hygroscoplus was deposited with the
Northern Utilization and Research Division, Agricultural
Research Service, U.S. Department of Agriculture,
Peoria, Illinois, U.S.A. under accession number NRRL
54 91. Rapamycin, and methods for its preparation by
culturing NRRL 54 91 are disclosed by U.S. Patent
3,929,992, issued December 30, 1975, the entire
disclosure of which is hereby incorporated by reference.
SUMMARY OF THE INVENTION
This invention relates to novel rapamycin
derivatives of the formula:
Formula II
30 wherein:
X is selected from the group consisting of H,
-OR 10 , -S(0) n R 10 , -NR 10 R 1:L , alkyl and aryl;
Y is selected from the group consisting of H,
-OR 10 , -S(0) n R 10 , -NR 10 R 11 , alkyl and aryl;
35 or X and Y taken together are =0;
n is selected from the group consisting of 0, 1
and 2 /
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R 1 is selected from the group consisting of =0,
(-OR 6 , H) and (H, H) ;
r2 is selected from the group consisting of =0,
(H,H), and (H, OH) /
r3 and R 6 are independently selected from the
group consisting of -H, C^-C^ alkyl, -C(=0)R 7 , -
C(=0)OR 7 ,
-C(=0)NHR 7 , and -C(=S)OR 7 ;
R 4 is selected from the group consisting of =0
and <H,OR 6 );
or R3 and R 4 can be taken together to form a
bridge of the formula A-C(R 8 ) (R 9 )-0-B, where A is a bond
to the oxygen bonded to the carbon at the 28-position
and B is a bond to the carbon at the 30-position;
R 5 is selected from the group consisting of -H
and C1-C4 alkyl;
R 7 is selected from the group consisting of C^-
Ciq alkyl, C3-C6 cycloalkyl, aryl groups, and
heterocyclic groups;
R 8 and R 9 are independently selected from the
group consisting of H, Ci to alkyl, or R 8 and R 9
taken together are =0;
R 1 ^ and R^- 1 are independently selected from the
group consisting of H, alkyl and aryl;
provided that,
(a) at least one of X and Y is H; and
(b) when Y is -R 10 , then R 10 is other than CH3;
and all pharmaceutical^ acceptable salts, hydrates or
solvates thereof.
This invention also relates to a pharmaceutical
composition comprising an effective amount of one or
more compounds of Formula II and a pharmaceutically
acceptable carrier or diluent.
This invention also relates to a method of
inhibiting the growth of pathogenic fungi in a human or
other animal in need thereof which comprises
administering an effective, non-toxic amount of one or
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more compounds of Formula II to such human or other
animal .
This invention also relates to a method of
inducing immunosuppression in a human or other animal in
5 need thereof which comprises administering an effective,
non-toxic amount of one or more compounds of Formula II
to such human or other animal .
In addition, this invention relates to a method
of treating carcinogenic tumors in a human or other
10 animal comprising administering an effective, non-toxic
amount of one or more compounds of Formula II to such
human or other animal .
Still further, this invention relates to a
method of preparing compounds of Formula II comprising
15 contacting a compound of the formula:
20
25
with an acid selected from the group consisting of
30 protic acids and Lewis acids, and with an appropriate
nucleophile .
DETAILED DESraTPTTOK OF THE INVENTION
When any substituent or variable (e.g., aryl,
35 alkoxyl, R 1 , R 2 , R 3 , R 5 , R 6 , etc.) occurs more than one
time in the formula of any of the compounds of Formula
II, such variable or substituent definition on each
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occurrence is independent of its definition at every
other occurrence, unless otherwise indicated.
Combinations of substituents and/or variables in a
constituent of the compounds of the invention are
permissible only if such combinations result in a stable
compound.
The parenthetical nomenclature used in the
definition of substituents such as R 1 (e.g., (H, OR 6 ) is
intended to reflect the substituents on both valences of
the relevant atom. The invention is not limited to
particular isomers and the order of moieties in the
parentheses does not suggest a particular configuration.
As used herein, except where otherwise noted,
the term "alkyl" is intended to include both branched-
and straight-chain, cyclic, acyclic and polycyclic,
saturated and unsaturated aliphatic hydrocarbon groups.
Preferably, alkyl groups have from one to six carbon
atoms, unless otherwise noted. Such alkyl group may be
optionally substituted by one or more members
independently selected from the group consisting of
aryl, hydroxyl, protected hydroxyl, C^Cq alkoxyl,
acyloxy, amino, N-acylamino, ketone, halogen, cyano, and
carboxyl . The term "alkyl" also includes the above-
mentioned groups in which a heteroatom selected from
oxygen, nitrogen and sulfur is substituted for one or
more carbon atoms in the alkyl moiety.
As used herein, the term "aryl" is intended to
include cyclic, heterocyclic, polycyclic and
heteropolycyclic unsaturated C4 to C14 moieties,
especially phenyl or naphthyl . Such aryl moieties may
be optionally substituted by one to five members
independently selected from the group consisting of C^-
Cg alkyl, aryl, hydroxyl, protected hydroxyl, Ci-Cg
alkoxyl, acyloxy, amino, N-acylamino, -S(0)n alkyl,
nitro, cyano, carboxyl and halogen.
As used herein, the term "alkoxyl" represents an
alkyl group as herein defined of the indicated number of
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carbon atoms attached through an oxygen bridge.
As used herein, the term "acyloxy" is intended
to represent the groups -0C (0) - (alkyl) and 0C (0) - (aryl) .
As used herein, the term "amino" is intended to
represent the groups -NH2, -N (alkyl) 2.1 -NH (alkyl), -
N (aryl) 2/ and -NH(aryl).
As used herein, the term "N-acylamino" is
intended to represent the groups -NHC (0) - (alkyl) and
-NHC (0) - (aryl) .
As used herein, the term ''ketone' 1 is intended to
mean the moiety -C(0)-.
As used herein, the term "halogen is intended to
include fluoro, chloro, bromo and iodo.
As used herein, the term "cycloalkyl" is
intended to include saturated and unsaturated
hydrocarbon aliphatic ring groups having the specified
number of carbon atoms . Such cycloalkyl may be
optionally substituted by one or more members
independently selected from the group consisting of
aryl, hydroxyl, Ci-Cg alkoxyl, acyloxy, amino, N-
acylamino, ketone, and halogen.
As used herein, the term "heterocycle" is
intended to include a stable 5- to 7-membered mono- or
bicyclic or stable 7- to 10-membered bicyclic
heterocyclic ring which is either saturated or
unsaturated, and which consists of carbon atoms and from
one to three heteroatoms independently selected from the
group consisting of N, 0 and S, and wherein the nitrogen
and sulfur heteroatoms may optionally be oxidized, and
the nitrogen heteroatom may optionally be quarternized,
and including any bicyclic group in which any of the
above-defined heterocyclic rings is fused to a benzene
ring. The heterocyclic ring may be attached at any
heteroatom or carbon atom which results in the creation
of a stable structure. Examples of such heterocyclic
elements include but are not limited to piperidyl,
piperidinyl, piperazinyl, pyrimidinyl, pyridazinyl,
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oxazolyl, furyl, and thienyl . The heterocycle may be
optionally substituted in a manner such that carbon
atoms attached to a heteroatom are not directly
substituted by a heteroatom, by from one to four members
independently selected from the group consisting of C]_-
Cg alkyl, aryl, hydroxyl, C^-Ce alkoxyl, acyloxy, amino,
N-acylamino, nitro and halogen.
Naturally occurring rapamycin has a structure in
which there is a methoxy group in the S-conf iguration
( OMe) at the C-7 atom of the molecule. Using the
novel synthetic methods disclosed herein, derivatives of
rapamycin have been prepared, including compounds
derivatized in both the R- (X is other than H) and S- (Y
is other than H) configurations at the C-7 atom as well
as the compound epirapamycin, in which there is a
methoxy group in the R-conf iguration ( OMe) at the C-
7 atom.
Preferred compounds of the invention include the
following compounds wherein, together or independently:
(1) R 1 is (H,OH)
(2) R 2 is =0
(3) R 3 is H
(4) R 4 is =0
(5) R 5 is H
(6) X and Y are each H
(7) One of X and Y is selected from the group
consisting of OR 10 and SR 10 or where X and Y are taken
together as =0
(8) One of X and Y is selected from the group
consisting of -OR 10 and -SR 10 where R 10 is selected from
the group consisting of optionally substituted C1-C3
alkyl, optionally substituted benzyl, and optionally
substituted phenyl
(9) One of X and Y is an alkyl group or when one
of R 10 and R 11 is an alkyl group, said alkyl is selected
from the group consisting of branched, straight-chain,
cyclic, polycyclic, saturated and unsaturated alkyl
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groups which may be substituted with 0, 1 or more
substitutents selected from the group consisting of
aryl, keto, hydroxyl, alkoxyl, acyloxy, amino, N-
acylamino, halogen, cyano and carboxyl substituents, and
5 in which at least one carbon atom may be replaced with a
heteroatom selected from the group consisting of O, S
and N
(10) One of X and Y is an aryl group or when one
of R 10 or R 11 is an aryl group, said aryl group is
10 selected from the group consisting of cyclic,
heterocyclic, polycyclic and heteropolycyclic C 2 to C14
aryl groups, which may be substituted by one to five
members selected from the group consisting of alkyl,
hydroxyl, alkyoxyl, acyloxy, amino, N-acylamino,
15 halogen, cyano, carboxyl and nitro groups.
Specifically preferred compounds of this
invention include those where:
(1) R 1 is <H,OH), R 2 is =0, R 3 is H, R 4 is =0,
R 5 is H, Y is H and X is -OCH3 .
20 (2) R 1 is (H,OH), R 2 is =0, R 3 is H, R 4 is =0,
R 5 is H, Y is H and X is -OC2H5 .
(3) R 1 is (H,0H), R 2 is =0, R 3 is H, R 4 is =0,
R 5 is H, X is H and Y is -OC 2 H 5 .
(4) R 1 is (H, OH), R 2 is =0, R 3 is H, R 4 is =0,
25 R 5 is H, Y is H and X is -OCH2CH2OH.
(5) R 1 is (H,0H), R 2 is =0, R 3 is H, R 4 is =0,
R 5 is H, X is H and Y is -0CH 2 CH 2 0H.
(6) R 1 is (H, OH), R 2 is =0, R 3 is H, R 4 is =0,
R 5 is H, Y is H and X is 3, 4-dimethoxybenzyloxy .
30 (7) R 1 is (H, OH), R 2 is =0, R 3 is H, R 4 is =0,
R 5 is H, X is H and Y is 3, 4-dimethoxybenzyloxy .
(8) R 1 is (H,0H), R 2 is =0, R 3 is H, R 4 is=0, R 5
is H, Y is H and X is -SCH3 .
(9) R 1 is (H,0H), R 2 is =0, R 3 is H, R 4 is=0, R 5
35 is H, X is H and Y is -SCH3 .
(10) R 1 is (H,OH), R 2 is =0, R 3 is H, R 4 is=0,
R 5 is H, Y is H and X is -SPh where Ph is phenyl.
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(11) R 1 is (H,0H), R 2 is =0, R 3 is H, R 4 is=0,
R 5 is H, X is H and Y is -SPh where Ph is phenyl.
(12) R 1 is (H,OH), R 2 is =0, R 3 is H, R 4 is=0,
R 5 is H, Y is H and X is 2-furanyl.
5 (13) R 1 is (H,OH), R 2 is =0, R 3 is H, R 4 is=0,
R 5 is H, X is H and Y is 2-furanyl.
(14) R 1 is (H,OH), R 2 is =0, R 3 is H, R 4 is=0,
R 5 is H, Y is H and X is O-acetyl .
(15) R 1 is (H,OH), R 2 is =0, R 3 is H, R 4 is=0,
10 r5 is H, and each of X and Y is H.
(16) R 1 is (H,0H), R 2 is =0, R 3 is H, R 4 is=0,
R 5 is H, and X and Y are taken together as =0.
(17) Rl is (H,0H), R 2 is =0, R 3 is H, R 4 is=0,
R 5 is H, X is H and Y is allyl.
15 (18) R 1 is (H,OH), R 2 is =0, R 3 is H, R 4 is=0,
R 5 is H, Y is H and X is OH.
(19) R 1 is (H,0H), R 2 is =0, R 3 is H, R 4 is=0,
R 5 is H, Y is H and X is 2 , 4-dimethoxyphenyl .
(20) Rl is (H,0H), R 2 is =0, R 3 is H, R 4 is=0,
20 R 5 is H, X is H and Y is 2 , 4-dimethoxyphenyl .
(21) R 1 is (H,0H), R 2 is =0, R 3 is H, R 4 is =0,
R 5 is H, X is H and Y is -OH.
The compounds of this invention can exist in
25 free form or, where appropriate, in salt form.
Pharmaceutically acceptable salts and their preparation
are well-known to those of skill in the art. The
pharmaceutically acceptable salts of the compounds of
this invention include the conventional non-toxic salts
30 or the quaternary ammonium salts of such compounds which
are formed, for example, from inorganic or organic acids
of bases .
The compounds of the invention may form hydrates
or solvates. It is known to those of skill in the art
35 that charged compounds form hydrated species when
lyophilized with water, or form solvated species when
concentrated in a solution with an appropriate organic
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solvent .
The compounds of this invention may be prepared
by the methods outlined below or conventional variations
thereof. The reagents utilized are either described in
the literature or are commercially available.
Compounds of Formula II (where X and Y are other
than =0) may be prepared from rapamycin or other
compounds of the formula:
III
where R 1 - R 11 and n are as defined above, by contacting
said compound with an acid selected from the group
consisting of protic acids and Lewis acids, and the
desired nucleophilic reagent, such as an alcohol, thiol,
electron-rich aromatic, allylsilane, etc. Preferred
acids are trif luoroacetic acid and titanium
tetrachloride, although these are by no means limiting
of the acids which may be utilized. The reaction is
preferably carried out in an inert atmosphere (e.g.,
under argon) at a temperature in the range of about -7 8°
to 0° C, preferably about -40° C. Suitable solvents
include nonprotic, nonpolar solvents, such as but not
limited to dichloromethane .
Compounds of the invention where X and Y are
taken together as =0 may be prepared from rapamycin or
other compounds of formula III by contacting said
compounds with a quinone oxidant such as DDQ (2,3-
dichloro-5, 6- dicyano-1 , 4-benzoquinone) .
'-10-
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Rapamycin derivatives reduced at the C-30
position may be prepared by treatment of rapamycin with
a mixture of cerium trichloride and sodium
cyanoborohydride . Suitable solvents for this reaction
5 include a mixture of acetic acid and tetrahydrof uran .
This reaction is illustrated in Scheme A with rapamycin
shown as the starting material; however other rapamycin
derivatives may be reduced at the C-30 position using
this method.
10 Scheme A
15
RAPAMYCIN
20
Compounds of the invention which are reduced at
both the C-14 and C-30 positions (i.e., R 2 and R 4 =
(H,OH) may be prepared by the action of
diisobutylaluminum hydride on rapamycin or a derivative
25 thereof. By appropriate control of the same reduction
method (e.g., low temperature, limiting hydride and
reaction times) , compounds reduced only at the C-14
position can be prepared. These reactions are
illustrated, with rapamycin shown as the starting
30 material, in Scheme B.
Scheme B
RAPAMYCIN
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Also illustrated in Scheme B is the preparation
of a C-13 O-methylated derivative, prepared by treatment
with acidic methanol.
Compounds of the invention having a bridge
5 between the C-28 and C-30 positions may be prepared by
methods analogous to that shown in Scheme C. The C-30
reduced derivative is contacted with a dialkoxypropane
such as dimethoxypropane to yield the desired compound
where R 8 and R 9 are each alkyl .
10
Scheme C
Another type of compound having a bridge between
the C-28 and C-30 positions may be prepared by methods
25 analogous to that shown in Scheme D. The C-30 reduced
derivative is contacted with carbonyldiimidazole to
yield the desired compound where R 8 and R 9 taken
together are =0.
Scheme D
30
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When the derivatives of this invention are
compounds derivatized at more than one atom of the
rapamycin molecule (e.g., at C-7, C-13, C-14, C-28, C-30
or C-43) , the synthetic order of the derivation can
5 vary. The preferred order of reactions will be
dependent upon the nature of the modifications at each
position and their compatibility with subsequent
reaction conditions, and would be obvious to chemists
skilled in the art.
10 The Examples provided below in this -
specification provide a variety of synthetic methods for
preparing compounds of this invention.
This invention also relates to a pharmaceutical
compositions comprising a pharmaceutically acceptable
15 carrier or diluent and an effective amount of one or
more compounds of Formula II.
A compound of Formula II is administered in
conventional dosage form prepared by combining a
therapeutically effective amount of the compound
20 ("active ingredient") with standard pharmaceutical
carrier or diluents according to conventional
procedures. These procedures may involve mixing,
granulating and compressing or dissolving the
ingredients as appropriate to the desired preparation.
25 The pharmaceutical carrier employed may be, for
example, either a solid or liquid. Exemplary of solid
carrier are lactose, terra alba, sucrose, talc, gelatin,
agar, pectin, acacia, magnesium stearate, stearic acid
and the like. Exemplary of liquid carriers are syrup,
30 peanut oil, olive oil, water and the like. Similarly,
the carrier or diluent may include time delay material
well known to the art, such as glyceryl monostearate or
glyceryl distearate along or with a wax, ethylcellulose,
hydroxypropylmethylcellulose, methylmethacrylate and the
35 like.
A wide variety of pharmaceutical forms can be
employed. Thus, if a solid carrier is used, the
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preparation can be tableted, placed in a hard gelatin
capsule in powder or pellet form or in the form of a
troche or lozenge. The amount of solid carrier will
vary widely but preferably will be from about 25 mg to
5 about 1 g. If a liquid carrier is used, the preparation
will be in the form of a syrup, emulsion, soft gelatin
capsule, sterile injectable solution or suspension in an
ampule or vial or nonaqueous liquid suspension.
To obtain a stable water soluble does form, a
10 pharmaceutically acceptable salt of a compounds of the
invention is dissolved in an aqueous solution of an
organic or inorganic acid, such as a 0 . 3M solution of
succinic acid, or, preferably, citric acid. If a
soluble salt form is not available, the compounds of the
15 invention is dissolved in a suitable cosolvent or
combinations thereof. Examples of such suitable
cosolvents include, but are not limited to, alcohol,
propylene glycol, polyethylene glycol 300, polysorbate
80, glycerin and the like in concentrations ranging from
20 0-60% of the total volume.
Tests indicate that the compounds of this
invention are useful in prophylactically or
therapeutically inhibiting the growth of pathogenic
fungi in a human or other animal in need thereof. The
25 invention, therefore, includes methods of inhibiting the
growth of pathogenic fungi in a human or other animal in
need thereof which comprises administering to such human
or animal an effective, non-toxic amount of a compound
of Formula II.
30 By the term "pathogenic fungi" is meant fungi
capable of producing disease in a human or other animal.
Examples of pathogenic fungi include, but are not
limited to Candida albicans and other Candida species,
Microsporum gypseum, Trichophyton mentagrophytes ,
35 Aspergillus sp . and Sporotrichum sp . The ability of the
compounds of this invention to inhibit the growth of
pathogenic fungi may be demonstrated or predicted by
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standard tests known and used for this purpose, for
example, the yeast assay described below.
One skilled in the art would be able, by routine
experimentation, to determine what an effective, non-
5 toxic amount of compound would be for the purpose of
inhibiting pathogenic fungi growth. Generally, however,
an effective dosage will be in the range of about 0.05
to 100 milligrams per kilogram body weight per day.
Tests indicate that the compounds of this
10 invention are also useful for inducing
immunosuppression, i.e., inducing a suppression of a
human's or animals immune system. This invention
therefore relates to a method of prophy lactically or
therapeutically inducing immunosuppression in a human or
15 other animal in need thereof which comprises
administering an effective, non-toxic amount of such a
compound of this invention to such human or other
animal .
The ability of the compounds of this invention
20 to induce immunosuppression may be demonstrated in
standard tests used for this purpose, for example, a
mixed lymphocyte reaction test or a test measuring
inhibition of T-cell proliferation measured by thimidine
uptake .
25 The fact that the compounds of this invention
have utility in inducing immunosuppression means that
they are useful in the treatment or prevention of
resistance to or rejection of transplanted organs or
tissues (e.g., kidney, heart, lung, bone marrow, skin,
30 cornea, etc.); the treatment or prevention of
autoimmune, inflammatory, proliferative and
hyperprolif erative diseases, and of cutaneous
manifestations of immunologically mediated diseases
(e.g., rheumatoid arthritis, lupus erythematosus,
35 systemic lupus erythematosus, Hashimotos thyroiditis,
multiple sclerosis, myasthenia gravis, type 1 diabetes,
uveitis, nephrotic syndrome, psoriasis, atopical
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dermatitis, contact dermatitis and further eczematous
dermatitides, seborrheic dermatitis, Lichen planus,
Pemplugus, bullous Pemphigoid, Epidermolysis bullosa,
uritcaris, angiodemas, vasculitides, erythemas,
5 cutaneous eosinophilias, Alopecia areata, etc.); the
treatment of reversible obstructive airways disease,
intestinal inflammations and allergies (e.g., Coeliac
disease, proctitis, eosinophilia gastroenteritis,
mastocytosis, Chrohn ' s disease and ulcerative colitis)
10 and food related allergies (e.g., migrane, rhinitis, and
eczema) .
One skilled in the art would be able, by routine
experimentation, to determine what an effective, non-
toxic amount of compound would be for the purpose of
15 inducing immunosuppression. Generally, however, an
effective dosage will be in the range of about 0.05 to
100 milligrams per kilogram body weight per day.
The compounds of this invention should also be
useful for treating carcinogenic tumors in a mammal.
20 More specifically, the compounds should be useful for
reducing tumor size, inhibiting tumor growth and/or
prolonging the survival time of tumor-bearing animals,
Accordingly, this invention also relates to a method of
treating carcinogenic tumors in a human or other animal
25 comprising administering to such human or animal an
effective, non-toxic amount of a compound of Formula II.
One skilled in the art would be able, by routine
experimentation, to determine what an effective, non-
toxic amount of compound would be for the purpose of
30 treating carcinogenic tumors. Generally, however, an
effective dosage is expected to be in the range of about
0.05 to 100 milligrams per kilogram body weight per day.
The compounds of the invention may be
administered to a human or other animal in accordance
35 with the aforementioned methods of treatment in an
amount sufficient to produce such effect to a
therapeutic or prophylactic degree. Such compound of the
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invention can be administered to such human or other
animal in a conventional dosage form prepared by
combining the compound of the invention with a
conventional pharmaceutically acceptable carrier or
5 diluent according to known techniques . It will be
recognized by one of skill in the art that the form and
character of the pharmaceutically acceptable carrier or
diluent is dictated by the amount of active ingredient
with which it is to be combined, the route of
10 administration and other well-known variables.
The route of administration of the compound of
the invention may be oral, parenteral, by inhalation or
topical. The term parenteral as used herein includes
intravenous, intramuscular, subcutaneous, rectal,
15 vaginal or intraperitoneal administration. The
subcutaneous and intramuscular forms of parenteral
administration are generally preferred.
The daily parenteral and oral dosage regimens
for employing compounds of the invention to
20 prophylactically or therapeutically inhibit the growth
of pathogenic fungi, to prophylatically or
therapeutically induce immunosuppression, or to
therapeutically treat carcinogenic tumors will generally
be in the range of about 0.05 to 100, but preferably
25 about 0.5 to 10, milligrams per kilogram body weight per
day .
The compounds of the invention may also be
administered by inhalation. By "inhalation" is meant
intranasal and oral inhalation administration.
30 Appropriate dosage forms for such administration, such
as an aerosol formulation or a metered dose inhaler, may
be prepared by conventional techniques. The preferred
dosage amount of a compound of the invention to be
employed is generally within the range of about 10 to
35 100 milligrams.
The compounds of the invention may also be
administered topically. By topical administration is
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meant non-systemic administration and includes the
application of a compounds of the invention externally
to the epidermis, to the buccal cavity and instillation
of such a compound into the ear, eye and nose, and where
5 the compound does not significantly enter the blood
stream. By systemic administration is meant oral,
intravenous, intraperitoneal and intramuscular
administration. The amount of a compound of the
invention (hereinafter referred to as the active
10 ingredient) required for therapeutic or prophylactic
effect on pathogenic fungi growth inhibition or
immunosuppression induction upon topical administration
will, of course, vary with the compound chosen, the
nature and severity of the condition being treated and
15 the animal undergoing treatment, and is ultimately at
the discretion of the physician. A suitable topical
dose of a compound of the invention will generally be
within the range of about 1 to 100 milligrams per
kilogram body weight daily.
20 While it is possible for an active ingredient to
be administered alone as the raw chemical, it is
preferable to present it as a pharmaceutical
formulation. The active ingredient may comprise, for
topical administration, from 0.001% to 10% w/w, e.g.,
25 from 1% to 2% by weight of the formulation although it
may comprise as much as 10% w/w but preferably not in
excess of 5% w/w and more preferably from 0.1% to 1% w/w
of the formulation.
The topical formulations of the present
30 invention, both for veterinary and for human medical
use, comprise an active ingredient together with one or
more acceptable carrier (s) therefore and optionally any
other therapeutic ingredient (s) . The carrier (s) must be
"acceptable" in the sense of being compatible with the
35 other ingredients of the formulation and no deleterious
to the recipient thereof.
Formulations suitable for topical administration
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include liquid or semi-liquid preparations suitable for
penetration through the skin to the site of where
treatment is required such as: liniments, lotions,
creams, ointments or pastes, and drops suitable for
5 administration to the eye, ear or nose.
Drops according to the present invention may
comprise sterile aqueous or oily solutions or
suspensions and may be prepared by dissolving the active
ingredient in a suitable aqueous solution of a
10 bactericidal and/or fungicidal agent and/or any other
suitable preservative, and preferably including a
surface active agent. The resulting solution may then be
clarified by filtration, transferred to a suitable
container which is then sealed and sterilized by
15 autoclaving or maintaining at 90-100 C for half an hour.
Alternatively, the solution may be sterilized by
filtration and transferred to the container by an
aseptic technique. Examples of bactericidal and
fungicidal agents suitable for inclusion in the drops
20 are phenylmercuric nitrate or acetate (0.002%),
benzalkonium chloride (0.01%) and chlorhexidine acetate
(0.01%). Suitable solvents for the preparation of an
oily solution include glycerol, diluted alcohol and
propylene glycol .
25 Lotions according to the present invention
include those suitable for application to the skin or
eye. An eye lotion may comprise a sterile aqueous
solution optionally containing a bactericide and may be
prepared by methods similar to those for the preparation
30 of drops. Lotions or liniments for application to the
skin may also include an agent to hasten drying and to
cool the skin, such as an alcohol or acetone, and/or a
moisturizer such as glycerol or an oil such as castor
oil or arachis oil .
35 Creams, ointments or pastes according to the
present invention are semi-solid formulations of the
active ingredient for external application. They may be
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made by mixing the active ingredient in finely-divided
or powdered form, alone or in solution or suspension in
an aqueous or non-aqueous fluid, with the aid of
suitable machinery, with a greasy or non-greasy basis.
5 The basis may comprise hydrocarbons such as hard, soft
or liquid paraffin, glycerol, beeswax, a metallic soap;
a mucilage; an oil of natural origin such as almond,
corn, arachis, castor or olive oil; wool fat or its
derivatives, or a fatty acid such as stearic or oleic
10 acid together with an alcohol such as propylene glycol
or macrogols. The formulation may incorporate any
suitable surface active agent such as an anionic,
cationic or non-ionic surface active such as sorbitan
esters or polyoxyethylene derivatives thereof.
15 Suspending agents such as natural gums, cellulose
derivatives or in organic materials such as silicaceous
silicas, and other ingredients such as lanolin, may also
be included.
It will be recognized by one of skill in the art
20 that the optimal quantity and spacing of individual
dosages of the compound of the invention will be
determined by the nature and extent of the condition
being treated, the form, route and site of
administration, and the particular animal being treated,
25 and that such optimums can be determined by conventional
techniques. It will also be appreciated by one of skill
in the art that the optimal course of treatment, i.e.,
the number of doses of the compound of the invention
given per day for a defined number of days, can be
30 ascertained by those skilled in the art using
conventional course of treatment determination tests.
Without further elaboration, it is believed that
one skilled in the art can, using the preceding
description, utilize the present invention to its
35 fullest extent. The following Examples are, therefore,
to be construed as merely illustrative and not a
limitation of the scope of the present invention in any
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way .
EXAMPLES
I. COMPOSITION EXAMPLES
EXAMPLE A - CAPSULE COMPOSITION
A pharmaceutical composition of this invention in
the form of a capsule is prepared by filling a standard
two-piece hard gelatin capsule with 50 mg of a compound
of the invention, in powdered form, 100 mg of lactose,
32 mg of talc and 8 mg of magnesium stearate .
EXAMPLE B - INJECTABLE PARENTERAL COMPOSITION
A pharmaceutical composition of this invention in
a form suitable for administration by injection is
prepared by stirring 1.5% by weight of a compound of the
invention in 10% by volume propylene glycol and water.
The solution is sterilized by filtration.
EXAMPLE C - OINTMENT COMPOSITION
Compound of the invention 1.0 g
White soft paraffin to 100.0 g
The compound of the invention is dispersed in a
small volume of the vehicle and gradually incorporated
into the bulk of the vehicle to produce a smooth,
homogeneous product. Collapsible metal tubes are then
filled with the dispersion.
EXAMPLE D - TOPICAL CREAM COMPOSITION
Compound of the invention 1.0 g
Polawax GP 200 20.0 g
Lanolin Anhydrous 2.0 g
White Beeswax 2.5 g
Methyl hydroxybenzoate 0.1 g
Distilled Water to 100.0 g
The polawax, beeswax and lanolin are heated
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together at 60 C. A solution of methyl hydroxybenzoate
is added and homo gen iz at ion is achieved using high speed
stirring. The temperature is then allowed to fall to 50
C. The compound of the invention is then added and
dispersed throughout, and the composition is allowed to
cool with slow speed stirring.
EXAMPLE E - TOPICAL LOTION COMPOSITION
Compound of the invention 1.0 g
Sorbitan Monolaurate 0.6 g
Polysorbate 20 0.6 g
Cetostearyl Alcohol 1.2 g
Glycerin 6.0 g
Methyl Hydroxybenzoate 0.2 g
Purified Water B.P. to 100.00 ml
The methyl hydroxybenzoate and glycerin are
dissolved in 70 ml of the water at 75 . The sorbitan
monolaurate, polysorbate 20 and cetostearyl alcohol are
melted together at 75 C and added to the aqueous
solution. The resulting emulsion is homogenized, allowed
to cool with continuous stirring and the compound of the
invention is added as a suspension in the remaining
water. The whole suspension is stirred until
homogeni zed .
EXAMPLE F - EYE DROP COMPOSITION
Compound of the invention 0.5 g
Methyl Hydroxybenzoate 0.01 g
Propyl Hydroxybenzoate 0.04 g
Purified water B.P. to 100.00 ml (B.P. =Brit ish
Pharmacopia)
The methyl and propyl hydroxybenzoates are
dissolved in 70 ml purified water at 75 C and the
resulting solution is allowed to cool . The compound of
the invention is then added, and the solution is
sterilized by filtration through a membrane filter (0.22
mu m pore size) and packed aseptically into suitable
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sterile containers.
EXAMPLE 6 - COMPOSITION FOR ADMINISTRATION BY
INHALATION
5 For an aerosol container with a capacity of 15-20
ml: Mix 10 mg of a compound of the invention with 0.2-
0.2% of a lubricating agent, such as polysorbate 85 or
oleic acid, and disperse such mixture in a propellant,
such as freon, preferably in a combination of (1,2
10 dichlorotetraf luoroethane) and dif luorochloromethane and
put into an appropriate aerosol container adapted for
either intranasal or oral inhalation administration
EXAMPLE H - COMPOSITION FOR ADMINISTRATION BY
15 INHALATION
For an aerosol container with a capacity of 15-20
ml: Dissolve 10 mg of a compound of the invention in
ethanol (6- 8 ml), add 0.1-0.2% of a lubricating agent,
such as polysorbate 85 or oleic acid; and disperse such
20 in a propellant, such as freon, preferably a combination
of (1.2 dichlorotetraf luoroethane) and
dif luorochloromethane, and put into an appropriate
aerosol container adapted for either intranasal or oral
inhalation administration.
II. SYNTHETIC EXAMPLES
In the following Examples, all starting materials
and chemical reagents were obtained from commercial
30 suppliers unless otherwise indicated. Rapamycin was
obtained from fermentation as described above.
General Procedure for C-7 Solvolysis
The following Methods A and B were used to
35 prepare compounds of Formula II.
Method A: Rapamycin (91.4mg, 0.10 mmol) in dry
dichloromethane (5mL) was treated with trif luoroacetic
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acid (0.5mL) at -4 0° C under argon. A bright yellow
color developed immediately and the resulting solution
was stirred at -40° C for 30 to 60 min . After that
time, an excess of the appropriate nucleophile was added
5 dropwise and stirring was continued for 30 min at -40°
C. The mixture was then partitioned between ethyl
acetate and saturated aqueous sodium bicarbonate.
Layers were separated and the organic layer successively
washed with saturated sodium bicarbonate, brine and
10 dried over anhydrous sodium sulfate. The resulting
crude material was subjected to preparative HPLC
purification to afford the desired 7-demethoxy-7-
substituted rapamycins .
Method B: Rapamycin (100 mg, 0.109 mmol) in dry
15 dichloromethane (2.5mL) was treated with trif luoroacetic
acid (lOOu L) at -40° C under argon. A bright yellow
color developed immediately and the resulting solution
was stirred at -40° C for 10 min. After that time, ten
to fifteen equivalents of the appropriate nucleophile
20 was added dropwise and stirring was continued for 15 min
at -4 0° C. The mixture was then partitioned between
dichloromethane and 5% aqueous sodium bicarbonate.
Layers were separated and the organic layer successively
washed with saturated sodium bicarbonate, brine and
25 dried over anhydrous sodium sulfate. The resulting
crude material was subjected to preparative HPLC
purification to afford the desired 7-demethoxy-7-
substituted rapamycins.
Example 1. 7-Demethoxy-7 (S) -ethoxyrapamycin
Method A was used to prepare the title compound:
1 H NMR(CDCl3, 400 MHz, 4:1 mixture of trans :cis amide
rotamers; data for the trans-rotamer) ; _ 6.375 (dd,
35 J=14.8, 10.5Hz, 1H) , 6.300 (dd, J=14.8, 9.8Hz, 1H) ,
6.134 (dd, J=14.9, 9.5Hz, 1H) , 5.940 (d, J=10.0 Hz, 1H) ,
5.533 (dd, J=14.9, 9.2Hz, 1H) , 5.408 (d, J=9.7 Hz, 1H) ,
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5.29 (m, 1H) , 5.17 (m, 1H) , 4.799 <s, 1H) , 4.173 (d,
J=5.9 Hz, 1H) , 3.87 (m, 1H) , 3.775 (dd, J=7.8, 7.0Hz,
1H) 3.714 (d, J=5.9 Hz, 1H) , 3.576 (br d, J=14.0 Hz,
1H) , 3.407 (s, 3H) , 3.338 (s, 3H) , 3.161 (dq, J=9.8,
5 6.9Hz, 1H) , 2.587 (dd, J=17.0, 6.3Hz, 1H) , 1.751 (s,
3H) , 1.662 (s, 3H) , 1.152 <t, J=6.9 Hz, 3H) , 1.098 (d,
J=6.7 Hz, 3H) , 1.047 (d, J=6.5 Hz, 3H) , 0.991 (d, J=6.6
Hz, 3H) , 0.951 (d, J=6.6 Hz, 3H) , 0.917 (d, J=6.7 Hz,
3H) , 0.668 (q, J=12 . 0 Hz, 1H) ; MS <ESl + /NH 4 OAc) m/z 950
10 (M+Na + ) , 945 (M+NH 4 + ) ; MS (ESl~/NH 4 COOH) m/z 972
(M+HCOO - ); UV (MeOH) max 267, 277, 289 nm.
Example 2. 7-Demethoxy-7 (R) -ethoxyrapamycin
Method A was used to prepare the title compound:
15 1 H NMR(CDCl3, 400 MHz, 2.5:1 mixture of transrcis amide
rotamers; data for the trans-rotamer ) : _ 6.365 (dd,
J=14.2, 11.0Hz, 1H) , 6.185 (dd, J=14.2, 10.4Hz, 1H) ,
6.12-6.05 (m, 2H) , 5.471 (dd, J=14.4, 9.1Hz, 1H) , 5.421
(d, J=10.3 Hz, 1H) , 5.25-5.20 (m, 2H) , 4.260 (br s, 1H) ,
20 4.10 - 4 . 04 (m, 1H) , 4.050 (s, 1H) , 4.012 (d, J=3 . 6 Hz,
1H) , 3.698 (dd, J=8.1, 1.5Hz, 1H) , 3.394 (s, 3H) , 3.332
(s, 3H) , 3.232 (dt, J=13.4, 7.0Hz, 1H) , 2.718 (dd,
J=17.4, 2.7Hz, 1H) , 2.376 (dd, J=17.4, 8.5Hz, 1H) , 1.750
<s, 3H) , 1.654 (s, 3H) , 1.180 (t, J=7 . 0 Hz, 3H) , 1.059
25 (d, J=6.7 Hz, 3H) , 1.005 (d, J=6.6 Hz, 3H) , 0.941 (d,
J=6.5 Hz, 3H) , 0.925 (d, J=6.5 Hz, 3H) , 0.863 (d, J=6.6
Hz, 3H) , 0.646 (q, J=12 . 0 Hz, 1H) ; MS (ES1 + /NH 4 OAc) m/z
950 (M+Na + ) , 945 (M+NH 4 + ) ; MS (ESl"/NH 4 COOH) m/z 972
(M+HCOO - ); UV (MeOH) max 267, 277, 289 nm.
30
Example 3 . 7-Demethoxy-7 (S) - (3 , 4-dimethoxybenzyl-
oxy) rapamycin
Method A was used to prepare the title compound:
3-H NMR(CDCl3, 400 MHz, 5:1 mixture of trans:cis amide
35 rotamers; data for the trans-rotamer); _ 6.87-6.79 (m,
3H) , 6.412 (dd, J=14.7, 10.9Hz, 1H) , 6.291 (dd, J=14.7,
10.4Hz, 1H) , 6.148 (dd, J=14.9, 10.4Hz, 1H) , 5.926 (d,
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J=10.9 Hz, 1H) , 5.503 (dd, J=14.9, 9.1Hz, 1H) , 5.394 (d,
J=9.9 Hz, 1H) , 5.211 (d, J=4.8 Hz, 1H) , 5.142 (td,
J=6.0, 4.0Hz, 1H) , 4.744 (s, 1H) , 4.418 (d, J=12.0 Hz,
1H) , 4.157 (d, J=6.0, Hz, 1H) , 4.110 (d, J=12 . 0 Hz, 1H) ,
5 3.884 (s, 3H) , 3.867 (s, 3H) , 3.824 (t, J=7 . 6 Hz, 1H) ,
3.502 (br d J=13.4 Hz, 1H) , 3.404 (s, 3H) , 3.325 (s,
3H) , 2.732 (dd, J=17.0, 5.8Hz, 1H) , 2.539 (dd, J=17.0,
6.3Hz, 1H) , 1.735 (s, 6H) , 1.081 (d, J=6.7 Hz, 3H) ,
1.054 (d, J=6.6 Hz, 3H) , 0.988 (d, J=6 . 6 Hz, 3H) , 0.931
10 (d, J=6.6 Hz, 3H) , 0.893 (d, J=6,7 Hz, 3H) , 0.661 (q,
J=11.9 Hz, 1H) / 13 C NMR (CDC1 3 , 100.6 MHz, 5:1 mixture
of trans :cis amide rotamers; data for the trans-
rot amer) : _ 215.7, 208.2, 192.7, 169.3, 166.7, 149.1,
148.5, 140,5, 136.0, 135.9, 133.3, 131.3, 130.4, 129.7,
15 126.8, 126.5, 119.8, 111.0, 110.7, 98.6, 84.9, 84.4,
81.3, 77.2, 75.5, 74.0, 69.3, 67.3, 59.5, 56.6, 55.9,
55.8, 51.4, 46.5, 44.3, 41.4; MS (ESl + /NH 4 OAc) m/z 1072
(M+Na + ) , 1049 (M+NH 4 + ) ; UV (MeOH) max 267, 278, 289 nm.
20 Example 4. 7-Demethoxy-7 (S) - (2-hydroxyethoxy) rapamycin
Method B was used to prepare the title compound:
1 H NMR(CDCl3, 400 MHz, 4:1 mixture of trans :cis amide
rotamers; data for the trans-rotamer) : _ 6.371 (dd,
J=14.6, 10.0Hz, 1H) , 6.303 (dd, J=14.6, 9.6Hz, 1H) ,
25 6.138 (dd, J=15.0, 9.6Hz, 1H) , 5.962 (d, J=9.7 Hz, 1H) ,
5.538 (dd, J=15.0, 8.9Hz, 1H) , 5.408 (d, J=9.9 Hz, 1H) ,
5.282 (d, J=4.7 Hz, 1H) , 5.164 (td, J=6.2, 4.0Hz, 1H) ,
4.849 (s, 1H) , 4.170 (d, J=6.0 Hz, 1H) , 3.822 (dd,
J=8.0, 7.2Hz, 1H) , 3.711 (d, J=6.0 Hz, 1H) , 3.683 (t,
30 J=4.0 Hz, 1H) , 3.582 (br d, J=14.0 Hz, 1H) , 3.408 (s,
3H) , 3.336 (s, 3H) , 3.244 (ddd, J=10.5, 5.5, 4.0Hz, 1H) ,
2.739 (dd, J=16.8, 6.0Hz, 1H) , 2.595 (dd, J=16.8, 6.3Hz,
1H) , 1.748 (s, 3H), 1.668 (s, 3H) , 1.103 (d, J=6.8 Hz,
3H) , 1.050 (d, J=6.7 Hz, 3H) , 0.994 (d, J=6.5 Hz, 3H) ,
35 0.950 (d, J=6.6 Hz, 3H) , 0.919 (d, J=6.7 Hz, 3H) , 0.668
(q, J=11.9 Hz, 1H) ; MS (ESl + /NH 4 OAc) m/z 961 (M+NH 4 + ) ,
882, 864; MS (ESl"/NH 4 COOH) m/z 988 (M+HCOO - ) ; UV (MeOH)
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max 267, 277, 289 nm.
Example 5. 7— Demeth.oxy-7 (R) - (2— hydroxy ethoxy) rapamycin
Method A was used to prepare the title compound:
5 1 H NMR(CDCl3, 400 MHz 6:1 mixture of trans :cis amide
rotamers; data for the trans-rotamer) :_ 6.359 (dd,
J=14.2, 11.1Hz, 1H) , 6.174 (dd, J=14.2, 10.7Hz, 1H) ,
6.099 (dd, J=14.5, 10.7Hz, 1H) , 6.055 (d, J=12.0 Hz,
1H) , 5.448 (d, J=10.0 Hz, 1H) , 5.377 (dd, J=14.5, 9.6Hz,
10 1H) , 5.34 (m, 1H) , 5.084 (dd, J=6.1, 2.7Hz, iH) , 4.295
(t, J=10.0 Hz, 1H) , 4.327 (d, J=4 . 0 Hz, 1H) , 4.065 (d,
J=4.0 Hz, 1H) , 4.026 (dd, J=10.8 2.3Hz, 1H) , 3.781 (br
d, J=12.8 Hz, 1H) , 3.70-3.64 (m, IH) , 3.541 (br d,
J=14.0 Hz, IH) , 3.465 (br d, J=11.5 Hz, IH) , 3.394 (s,
15 3H) , 3.342 (s, 3H) , 3.234 (dd, J=9.9, 6.7Hz, IH) , 3.092
(br t, J=9.2 Hz, IH) , 2.715 (dd, J=17.8, 5.7Hz, IH) ,
2.560 (dd, J=17.8, 6.8Hz, IH) , 1.833 (s, 3H) , 1.677(8,
3H) , 1.049 (d, J=6.6 Hz, 3H) , 0.979 (d, J=6.5 Hz, 3H) ,
0.963 (d, J=6.5 Hz, 3H) , 0.932 (d, J=6.5 Hz, 3H) , 0.883
20 (d, J=6.8 Hz 3H) , 0.591 (q, J=11.9 Hz, IH) / MS
(ESl + /NH 4 OAc) m/z 966 (M+Na + ) , 961 (M+NH 4 + ) , 882; MS
(ESl"/NH 4 COOH) m/z 988
(M+HCOO - ) ; UV (MeOH) max 267, 277, 289 nm.
25 Example 6. 7-Demethoxy-7 (R) -acetoxyrapamycin
Method A was used to prepare the title compound:
!h NMR(CDCl3, 400 MHz, 4:1 mixture of trans :cis amide
rotamers; data for the trans-rotamer): _ 6.337 (dd,
J=14.6, 10.7Hz, IH) , 6.233 (dd, J=14.6, 10.5Hz, IH) ,
30 6.097 (d, J=10.7 Hz, IH) , 6.086(dd, J=14.9, 10.5Hz, IH) ,
5.459 (dd, J=14.9, 9.0Hz, IH) , 5.423(d, J=9.7 Hz, IH) ,
5.331 (d, J=10.3 Hz, IH) , 5.23-5.20 (m, 2H) , 4.270 (d,
J=3.7 Hz, IH) , 4.014 (d, J=3.7 Hz, IH) , 3.897 (t,
J=10.0 Hz, IH) , 3.562 (br d, J=14.5 Hz, IH) , 3.392 (s,
35 3H) , 3.333 (2, 3H) , 2.708 (dd, J=17.5, 3.8Hz, IH) , 2.405
(dd, J=17.5, 8.0Hz, IH) , 2.067 (s, 3H) , 1.751 (s, 3H) ,
1.727 (s, 3H) , 1.062 (d, J=6.5 Hz, 3H) , 1.018 (d, J=6.6
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Hz, 3H) , 0.952 (d, J=6.4 Hz, 3H) , 0.906 (d, J=6.5 Hz,
3H) , 0.873 (d, J=6.7 Hz, 3H) , 0.642 (q, J=11.8 Hz, 1H) /
13c NMR (CDCI3, 3:1 mixture of trans :cis amide rotamers;
data for trans-rotamer ) _ 212.4, 207.5, 195.5, 169.7,
5 166.2, 139.1, 137.7, 131.1, 130.4, 128.2, 125.9, 124.4,
98.7, 84.4, 84.0, 74.6, 73.9, 69.6; MS (FAB/NaCL) m/z
964 (M+Na + ); UV (MeOH) max 267, 277, 289 nm.
Example 7. 7-Demethoxy-7 (S) - (2 1 -furanyl) rapamycixi
10 Method A was used to prepare the title compound:
1 H NMR(CDCl3, 400 MHz, 3:1 mixture of trans :cis amide
rotamers; data for the trans-rotamer); 7.290 (d, J=l . 6
Hz, 1H) , 6.289 (dd, J=3.2, 1.6Hz, 1H) , 6.006 (d, J=3.2
Hz, 1H) , 5.615 (dd, J=15.1, 8.2Hz,. 1H), 5.424 (d, J=9.8
15 Hz, 1H) , 5.336 (d, J=4 . 9 Hz, 1H) , 5.191 (q, J=5 . 5 Hz,
1H) , 5.096 (1H, s), 4.200 (d, J=6.0 Hz, 1H) , 3.734 (d,
J=6.0 Hz, 1H) , 3.697 (dd, J=8.5, 8.1Hz, 1H) , 3.408 (S,
3H) , 3.357 (s, 3H) , 2.689 (d, J=6.2 Hz, 2H) , 1.775 (s,
3H) , 1.559 (s, 3H) , 1.136 (d, J=6.8 Hz, 3H) , 1.051 (d,
20 J=6.6 Hz, 3H) , 1.006(d, J=6.5 Hz, 3H) , 0.990 (d, J=6.5
Hz, 3H) , 0.929 (d, J=6.8 Hz, 3H) , 0.669 (q, J=12 . 0 Hz,
1H) ; MS (ESl + /NH 4 OAc) m/z 967 (M+NH 4 + ) , 932; MS (ESI"
/NH 4 COOH) m/z 994 (M+HCOO - ) ; UV (MeOH) max 268, 278, 290
nm .
25
Example 8. 7-Demethoxyrapamycin
Method B (trimethylsilane as the nucleophile) was
used to prepare the title compound: ^-H NMR(CDCl3, 400
MHz, 3:1 mixture of trans :cis amide rotamers; data for
30 the trans-rotamer): _ 6.353 (dd, J=14.0, 11.1Hz, 1H) ,
6.151 (dd, J=14.1, 10.5Hz, 1H) , 6.075 (dd, J=14.1,
10.4Hz, 1H) , 5.874 (d, J=11.0 Hz, 1H) , 5.387 (d, J=10.5
Hz, 1H) , 5.343 (dd, J=14.4, 10.5Hz, 1H) , 5.24-5.18 (m,
2H) , 4.248 (br t, 1H) , 4.197 (s, 1H) , 4.005 (d, J=4 . 0
35 Hz, 1H) , 3.91-3.87 (m, 1H) , 3.457 (br d, J=11.5 Hz, 1H) ,
3.397 (s, 3H) , 3.336 (s, 3H) , 2.719 (dd, J=17.6, 4.6 Hz,
1H), 2.445(dd, J=17.6, 7.4 Hz, 1H) , 1.793 (s, 3H) , 1.722
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(s, 3H) / 1.0535 (d, J=6.6 Hz, 3H) , 0.997 (d, J=6.7 Hz,
3H) , 0.940<d, J=6.5 Hz, 3H) , 0.913(d, J=6.7 Hz,. 3H) ,
0.891 (d, J=6.9 Hz, 3H) , 0.626 (q, J=11.9 Hz, 1H) ; 13 C
NMR (CDCI3, 3:1 mixture of trans :cis amide rotamers;
5 data for trans-rotamer) _ 212.4, 207.5, 195.5, 169.7,
166.2, 139.1, 137.7, 131.1, 130.4, 128.2, 125.9, 124.4,
98.7, 84.4, 84.0, 74.6, 73.9, 69.6; MS (ESl + /NH 4 OAc) m/z
906 (M+Na + ) , 901(M+NH 4 + ); MS (ESI - /NH4COOH) m/z 928
(M+HCOO - ); UV (MeOH) max 267, 277, 289 nm.
10
Example 9. 7-Demethoxy-7 (S) -methylthiorapamycin
Method B was used to prepare the title compound:
l-H NMR(CDCl3, 400 MHz, 3:1 mixture of transrcis amide
rotamer; data for the trans-rotamer) : _ 6.363 (dd,
15 J=15.4, 10.6Hz, 1H) , 6.32-6.09 (m, 2H) , 5.816 (d, J=10.5
Hz, 1H) , 5.537 (dd, J=15.0, 9.4Hz, 1H) , 5.403 (d, J=9.8
Hz, 1H) , 5.289 (d, J=5 . 7 Hz, 1H) , 5.20-5.14 (m, 2H) ,
5.020 (br s, 1H) , 4.158 (d, J=7.0, 1H) , 3.88-3.76 (m,
1H) , 3.617 (d, J=7.0 Hz, 1H) , 3.59-3.45 (m, 2H) , 3.407
20 (s, 3H) , 3.341 (s, 3H) , 2.99-2.90 (m, 1H) , 2.780 (dd,
J=16.9, 6.5Hz, 1H) , 2.641 (dd, J=16.9, 6.5Hz, 1H) , 1.883
(s, 3H) , 1.739 (s, 3H) , 1.720 (s, 3H) , 1.128 (d, J=6.8
Hz, 3H) , 1.156 (d, J=6.5 Hz, 3H) , 1.015 (d, J=6 . 5 Hz,
3H) , 0.966 (d, J=6.5 Hz, 3H) , 0.932 (d, J=6.7 Hz, 3H) ,
25 0.667 (q, J=11.4 Hz, 1H) / 13 C NMR (CDCI3, 3:1 mixture of
trans :cis amide rotamers; data for trans-rotamer)
215.7, 208.4, 190.7, 169.2, 166.9, 140.3, 136.3, 134.3,
133.3, 130.1, 129.4, 127.1, 126.4, 98.3, 85.0, 84.4,
75.9, 73.9, 67.4; MS (FAB/NaCl ) m/z 952 (M+Na + ) ; MS
30 <ESl + /NH 4 OAc) m/z 952 (M+Na + ) , 947, (M+NH 4 + ) , 882, 864;
MS (ESl - /NH 4 COOH) m/z 974 (M+HCOO - ); UV (MeOH) max 266,
277, 288 nm.
Example 10. 7 -Demethoxy-7 (R) -methylthiorapamycin
35 Method B was used to prepare the title compound:
1 H NMR(CDCl3, 400 MHz, 2.5:1 mixture of trans:cis amide
rotamers; data for the trans-rotamer) : _ 6.388 (dd,
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J=14.5, 11.0Hz, 1H) , 6.22-6.06 (m, 2H) , 5.884 (d, J=10.9
Hz, 1H) , 5.45-5.38 (m, 2H) , 5.22-5.14 <m, 2H) , 4.278 (br
t, 1H) , 4.065 (d, J=3.4 Hz, 1H) , 4.030 (s, 1H) , 3.48-
3.41 (m, 1H) 3.395 (s, 3H) , 3.334 (s, 3H) , 3.18-3.09 (br
5 t, J=9.8, 1H) , 2.710 (dd, J=17.5, 3.2Hz, 1H) , 2.384 (dd,
J=17.6, 7.8Hz, 1H) , 1.814 (s, 6H) , 1.754 (s, 3H) , 1.715
(s, 3H) , 1.059 (d, J=6.5 Hz, 3H) , 0.989 (d, J=6.5 Hz,
3H) , 0.939 (d, J=6.5 Hz, 3H) , 0.915 (d, J=6.8 Hz, 3H) ,
0.877 (d, J=6.9 Hz, 3H) , 0.636 (q, J=11.7 Hz, 1H) ; MS
10 (FAB/NaCl) m/z 952 (M+Na + ) ; MS (ESl + /NH 4 OAc) m/z 952
(M+Na + ) , 947 (M+NH 4 + ) , 882, 864; MS (ESl~/NH 4 COOH) m/z
974 (M+HCOO - ); UV (MeOH) max 266, 277, 288 nm.
Example 11. 7-Demethoxy-7 (S) -phenylthiorapamycin
15 Method B was used to prepare the title compound:
!h NMR(CDCl3, 400 MHz, 4:1 mixture of trans :cis amide
rotamers; data for the trans-rotamer) : _ 7.35-7.15 (m,
5H) , 6.300 (dd, J=14.4, 11.2Hz, 1H) , 6.17-6.06 (m, 2H) ,
5.552 (d, J=ll.l Hz, 1H) , 5.510 (dd, J=14.0, 8.1Hz, 1H) ,
20 5.378 (d, J=9.8 Hz, 1H) , 5.19-5.09 (m, 2H) , 4.111 (d,
J=7.2 Hz, 1H) , 3.824 (br t, 1H) , 3.711 (dd, J=12.0,
4.9Hz, 1H) , 3.549 (d, J=7.2 Hz, 1H) , 3.537 (br d, 1H) ,
3.409 (s, 3H) , 3.307 (s, 3H) , 2.99- 2.90 (m, 1H) , 2.785
(dd, J=17.0, 7.2Hz, 1H) , 2.611 (dd, J=16.9, 5.5Hz, 1H) ,
25 1.814 (s, 3H) , 1.707 (s, 3H) , 1.131 (d, J=6.7 Hz, 3H) ,
1.033 (d, J=6.5 Hz, 3H) , 0.994 (d, J=6.5 Hz, 3H) , 0.950
(d, J=6.5 Hz, 3H) , 0.921 (d, J=6.8, 3H) , 0.669 (q,
J=15.8 Hz, 1H) ; 13 C NMR (CDCI3, 4:1 mixture of trans :cis
amide rotamers, data for trans-rotamer) _ 214.8, 208.7,
30 193.1, 169.0, 166.4, 140.5, 136.5, 133.4, 132.3, 130.4,
128.5, 126.4, 125.4, 98.4, 85.2, 84.4, 75.9, 73.9, 67.5;
MS (ESl + /NH 4 OAc) m/z 1009 <M+NH 4 + ) ; MS <ESl"/NH 4 COOH)
m/z 1036 (M+HCOO - ); UV (MeOH) max 267, 277, 289 nm.
35 Example 12. 7-Demethoxy-7 (R) -phenylthiorapamycin
Method B was used to prepare the title compound:
1 NMR(CDCl3, 400 MHz, 4:1 mixture of trans :cis amide
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rotamers; data for the trans-rotamer) : _ 7.38-7.18 (m,
5H) , 6.37-6.28 (m, 14.2, 1H) , 6.13-5.89 (m, 2H) , 5.644
<d, J=10.8 Hz, 1H) , 5.421 (d, J=10.3 Hz, 1H) , 5.25-5.20
(m, 2H) , 4.260 (br s, 1H) , 5.40-5.28 (m, 2H) , 4.385 (br
5 t, J=11.0, 1H) , 4.226 (br t, 1H) , 4.078 (d, J=3.3, 1H) ,
3.563 (dd, J=10.7, 2.3Hz, 1H) , 3.386 (s, 3H) , 3.323 (s,
3H) , 3.200 (dd, J=10.5, 6.6Hz, 1H) , 3.16-3.06 (m, 1H) ,
2.98-2.87 (m, 2H) , 2.698 (dd, J=17.6, 3.0Hz, 1H) , 2.263
(dd, J=17.6, 8.4Hz, 1H) , 1.898 (s, 3H) , 1.770 (s, 3H) ,
10 1.062 (d, J=6.6 Hz, 3H) , 0.971 (d, J=6 . 6 Hz, 3H) , 0.926
(d, J=6.4 Hz, 3H) , 0.883 (d, J=6.5 Hz, 3H) , 0.845 (d,
J=6.8 Hz, 3H) , 0.623 (q, J=12.0 Hz, 1H) ; MS
(ESl + /NH 4 OAc) m/z 1009 (M+NH 4 + ) ; MS (ESI - /NH 4 COOH) m/z
1036 (M+HCOO - ) ; UV (MeOH) max 267, 277, 289 nm
15
Example 13. 7-Demethoxy-7 (S) -allylrapamycin
Method B; ^-H NMR(CDC1 3 , 400 MHz, 3:1 mixture of
trans :cis amide rotamers; data for the trans-rotamer): _
6.368 (dd, J=15.6, 11.4 Hz, 1H) , 6.257 (dd, J=15.4, 11.2
20 Hz, 1H) , 6.133 (dd, J=15.7, 11.1 Hz, 1H) , 5.785 (d,
J=11.4 Hz, 1H) , 5.69-5.60 (m, 1H) , 5.564 (dd, J=15.9,
9.4 Hz, 1H) , 5.413 (d, J=10.7 Hz, 1H) , 5.294 (d, J=6.2
Hz, 1H) , 5.21- 5.16 (m, 1H) , 5.02-4.87 (m, 2H) , 4.168
(d, J=7.2 Hz, 1H) , 3.774 (br . t, J=11.4, Hz, 1H) , 3.642
25 (d, J=7.2 Hz, 1H) , 3.562 (s, 1H) , 3.502 (br d J=13.4 Hz,
1H) , 3.410 (s, 3H) , 3.350 (s, 3H) , 2.99-2.86 (m, 2H) ,
2.755 (dd, J = 17.4, 7.0, 1H) , 2.663 (dd, J 17.4, 6.2,
1H) , 1.761 (s, 3H) , 1.607 (s, 3H) , 1.135 (d, J=6.7 Hz,
3H) , 1.044 (d, J=6.6 Hz, 3H) , 1.013 (d, J=6.6 Hz, 3H) ,
30 0. 972 (d, J=6.6 Hz, 3H) , 0.924 (d, J=6.8 Hz, 3H) , 0.671
(q, J=12.0 Hz, 1H) ; 13 C NMR (CDCI3, 3:1 mixture of
trans :cis amide rotamers; data for the trans- rotamer) _
215.6, 208.7, 192.0, 169.4, 167.1, 139.1, 137.6, 137.2,
136.4, 132.1, 130.1, 128.4, 127.2, 126.9, 115.3, 98.2,
35 84.8, 84.4, 77.7, 76.0, 73.9, 67.2;, MS (ESl + /NH 4 OAc)
m/z 946 (M+Na + ) , 941 (M+NH 4 + ) ; UV (MeOH) max 267, 278,
289 nm.
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Example 14. 7-Demethoxy-7 (R) - (2 ' -furanyl) rapamycin
Method A was used to prepare the title compound:
1 H NMR (CDCI3 / 400MHz, 3.5:1 mixture of trans :cis amide
5 rotamers; data for the trans-rotamer) : _ 7.302 (d,
J=1.8 Hz, 1H) , 6.356 (dd, J=14.6, 10.9Hz, 1H) , 6.292
(dd, J=3.0, 1.8Hz, 1H), 6.190 (dd, J=14.6, 10.4Hz, 1H) ,
6.044 (br d, J=10.9 Hz, 1H) , 6.019 (d, J=3 . 0 Hz, 1H) ,
5.390 (d, J=10.3 Hz, 1H) , 5.327 (dd, J=14.7, 9.4Hz, 1H) ,
10 5.24-5.09 (m, 2H) , 4.294 (br s) , 4.116 (d, J=3 . 1 Hz,
1H) , 3.805 (s, 1H) , 3.524 (br d, J=15 . 0 Hz, 1H) , 3.469
(dd, J=11.7, 3.6Hz, 1H), 3.391 (s, 3H) , 3.335 (s, 3H) ,
3.245 (dq, J=10.3, 6.6 Hz,lH), 2.673 (dd, J=17.7, 2.8
Hz, 1H) , 2.360 (dd, J=17.7, 8.4Hz, 1H) , 1.825 (s, 3H) ,
15 1.662 (s, 3H) , 1.058 (d, J=6.4 Hz, 3H) , 0.973 (d, J=6.5
Hz, 3H) , 0.924 (d, J=6 . 4 Hz, 3H) , 0.888 (d, J=6.4 Hz,
3H) , 0.883 (d, J=6.5 Hz, 3H) , 0.633 (q, J=11.6 Hz, 1H) ;
MS (ESI+/NH4OAC) m/z 972 (M+Na + ) , 967 (M+NH 4 + ) ; MS (ES1~
/NH 4 COOH) m/z 994 (M+HCOO") ; UV (MeOH) max 268, 278, 290
20 nm.
Example 15 . 7-Demethoxy-7 (S) - (2 • 4 ' -dimethoxyphenyl) -
rapamycin
Method A was used to prepare the title compound:
25 1 H NMR (CDCI3, 400MHz, 2.5:1 mixture of trans :cis amide
rotamers; data for the trans-rotamer): _ 7.087 (d,
J=8.4 Hz, 1H) , 6.450 (dd, J=8.4, 2.2Hz, 1H) , 6.422 (d,
J=2.2 Hz, 1H) , 6.326 (dd, J=14.6, 10.5Hz, 1H) , 6.232
(dd, J=14.6, 10.4Hz, 1H) , 6.14-6.07 (m, 2H) , 5.465 (dd,
30 J=14.9, 9.8Hz, 1H) , 5.421 (d, J=10.5 Hz, 1H) , 5.312 (d,
J=5.3 Hz, 1H) , 5.177 (q, J=5 . 4 Hz, 1H) , 4.202 (br d,
J=5.0 Hz, 1H) , 3.912 (t, J=8.4 Hz, 1H) , 3.792 (s, 3H) ,
3.738 (s, 3H) , 3.565 (br d, J=13.4 Hz, 1H) , 3.410 (s,
3H), 3.353 (s, 3H) , 2.772 (dd, J=17.1, 5.5Hz, 1H) , 2.596
35 (dd, J=17.1, 6.4 Hz, 1H) , 1.817 (s, 3H) , 1.500 (s, 3H) ,
1.096 (d, J=6.7 Hz, 3H), 1.038 (d, J=6.7 Hz, 3H) , 0.980
(d, J=6.5 Hz, 3H) , 0.935 (d, J=6.5 Hz, 3H) , 0.924 (d,
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J=6.7 Hz, 3H) , 0.674 (q, J=11.8 Hz, 1H) ; MS
(ESI+/NH4OAC) m/ z ' 1037 (M+NH 4 + ) ; MS (ESl~/NH 4 COOH) m / z
1064
(M+HCOO - ) .
5
Example 16 . 7-Demethoxy-7 (R) - (2 ' , 4 ' -dimethoxyphenyl) -
rapamycin
Method A was used to prepare the title compound:
1 H NMR (CDC13, 400MHz, 10:1 mixture of trans :cis amide
10 rotamers; data for the trans-rotamer) : 6.921 <d,
J=9.1 Hz, 1H) , 6.47-6.42 (m, 2H) , 6.371 (dd, J=14.7,
11.0Hz, 1H) , 6.225 (dd, J=14.7, 10.5Hz, 1H) , 6.090 (br
d, J=11.0 Hz, 1H) , 6.059 (dd, J=14.9, 10.5Hz, 1H) , 5.369
(d, J=10.4 Hz, 1H) , 5.29-5.23 (m, 2H) , 5.191 (br d,
15 J=2.8 Hz, 1H) , 4.323 (br s) , 4.205 (d, J=2 . 7 Hz, 1H) ,
3.906 (br d, J=11.4 Hz, 1H) , 3.790 (s, 3H) , 3.743 (s,
3H) , 3.579 (d, J=1.0 Hz, 1H) , 3.408 (br t, J-10.0 Hz,
1H) , 3.396 (s, 3H) , 3.341 (s, 3H) , 3.246 (dq, J=10.3,
6.4Hz, 1H) , 2.775 (d, J=18.0 Hz, 1H) , 2.274 (dd, J=18.0,
20 9.1Hz, 1H) , 1.884 (s, 3H) , 1.511 (s, 3H) , 1.069 (d,
J=6.5 Hz, 3H) , 0.926 (d, J=6 . 6 Hz, 3H) , 0.906 (d, J=6.5
Hz, 3H) , 0.890 (d, J=6.5 Hz, 3H) , 0.857 (d, J=6.6 Hz,
3H) , 0.651 (q, J=11.8 Hz, 1H) ; MS (ESl + /NH 4 OAc) m / z 1037
(M+NH 4 +) ; 1019 (M+Na + ); MS (ESl~/NH 4 COOH) m / z 1064
25 (M+HCOO - ) .
Example 17 . 7-Demethoxy-7-oxorapamycin
To a solution of rapamycin (5 mg, 0.055 mmol) in
dichloromethane (0.1 mL) was added water (0.02 mL)
30 followed by DDQ (2.4 mg, 0.011 mmol), and the resulting
dark brown suspension was stirred at room temperature
for 45 min. The mixture was filtered through celite and
then partitioned between dichloromethane and brine; the
organic extracts were dried over anhydrous magnesium
35 sulfate and the resulting crude material was purified by
prep TLC. The desired product was isolated as a
colorless oil (1.2 mg) . 1 E NMR (CDCI3, 400MHz, 3:1
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mixture of trans :cis amide rotamers; data for the trans-
rotamer) : _ 7.145 (d, J=10.4 Hz, 1H) , 6.635 (dd,
J-14.9, 10.6Hz, 1H) , 6.57-6.48 (m, 2H) , 6.224 (dd,
J=15.1, 10.6Hz, 1H) , 5.662 (dd, J=15.1, 8.6Hz, 1H) ,
5 5.447 (d, J=11.2 Hz, 1H) , 4.305 (br d, J=5 . 4 Hz, 1H) ,
3.983 (d, J=5.4 Hz, 1H) , 3.385 (s, 3H) , 3.346 (s, 3H) , ;
MS (ESl+/NH 4 OAc) m / z 920 (M+Na + ) , 915 (M+NH 4 + ) , 898,
880; UV (MeOH) max 318nm.
10 Example 18. 7-O-Demethylrapamycin
Method A was used to prepare the title compound:
1 H NMR(CDCl3, 400 MHz, 2.5:1 mixture of trans :cis amide
rotamers; data for the trans-rotamer) : 6.356 (dd,
J=14.8, 11.2Hz, 1H) , 6.241 (dd, J=14.8, 10.6Hz, 1H) ,
15 6.089 (dd, J=14.9, 10.6Hz, 1H) , 5.382 (dd, J=14.9,
9.7Hz, 1H) , 5.337 (d, J=11.0 Hz, 1H) , 5.28-5.21 (m, 2H) ,
4.580 (s, 1H) , 4.239 (br s, 1H) , 4.07-4.03 (m, 1H) ,
4.001 (d, J=4.0 Hz, 1H) , 3.539 (br d, J=13.0 Hz, 1H) ,
3.380 (s, 3H) , 3.333 (s, 3H) , 3.192 (d, J=2 . 8 Hz, 1H) ,
20 2.563 (dd, J=17.7, 5.0Hz, 1H) , 1.819 (s, 3H) , 1.684 (s,
3H) , 1.054 (d, J=6.6 Hz, 3H) , 0.991 (d, J=6.6 Hz, 3H) ,
0.970 (d, J=6.6 Hz, 3H) , 0.934 (d, J=6.4 Hz, 3H) , 0.879
(d, J=6.8 Hz, 3H) , 0.552 (q, J=11.8 Hz, 1H) ; MS
(FAB/NaCl) m/z 922 (M+Na + ) , MS (ESl + /NH 4 OAc) m/z 922
25 (M+Na + ) 917 (M+NH 4 + ), 882, 864; MS (ESl~/NH4COOH) m/z 944
(M+HCOO - ) ; UV (MeOH) max 266, 277, 288nm
Example 19. 7-0-Demethyl-7-epirapamycin
Method A was used to prepare the title compound:
30 1 H NMR(CDCl3, 400 MHz, 4:1 mixture of trans :cis amide
rotamers; data for the trans-rotamer): _ 6.397 (dd,
J=14.6, 10.9Hz, 1H) , 6.270 (dd, J=14.6, 10.7Hz, 1H) ,
6.160 (d, J=10.9 Hz, 1H) , 6.131 (dd, J=15.0, 10.7Hz,
1H) , 5.470 (dd, J=15.0, 8.7Hz, 1H) , 5.401 (d, J=9.8 Hz,
35 1H) , 5.271 (d, J=4 . 8 Hz, 1H) , 5.193 (td, J=6.4, 3.5Hz,
1H) , 4.799 (s, 1H) , 4.200 (br d, J=5 . 0 Hz, 1H) , 3.816
(d, J=5.4 Hz, 1H) , 3.573 (br d, J=13.0 Hz, 1H) , 3.396
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(s, 3H) , 3.337 (s, 3H) , 3.251 (d, J=2 . 5 Hz, IE), 2.729
(dd, J=17.4, 6.4Hz, 1H) , 2.631 (dd, J=17.4, 6.2Hz, 1H) ,
1.765 (s, 3H) , 1.715 (s, 3H) , 1.076 (d, J=6.8 Hz, 3H) ,
1.055 (d, J=6.7 Hz, 3H) , 0.978 (d, J=6.4 Hz, 3H) , 0.948
5 (d, J=6.6 Hz, 3H) , 0.914 (d, J=6.8 Hz, 3H) , 0.627 (q,
J=11.7 Hz, 1H) ; MS (FAB/NaCl) m/z 922 (M+Na + ) ; MS
(ESl + /NH 4 OAc) m/z 922 (M+Na4+) , 917 (M+NH 4 + ) , 882, 864;
MS (ESl"/NH 4 COOH) m/z 944 (M+HCOCT) / UV (MeOH) max 266,
277, 288 nm.
10
Example 20. 7-Epirapamycin
Method A was used to prepare the title compound:
1 H NMR(CDCl3, 400 MHz, 3:1 mixture of trans:cis amide
rotamers; data for the trans-rotamer) :_ 6.377 (dd,
15 J=14.4, 11.1Hz, 1H) , 6.23-6.07 (3H, m) , 5.481 (dd,
J=14.9, 8.7Hz, 1H) , 5.425 (d, J=10.2 Hz, 1H) , 5.24-5.20
(m, 2H) , 4.258 (br s, 1H) , 4.076 (d, J=0.9 Hz, 1H) ,
4.005 (d, J=4.7 Hz, 1H) , 3.618 (dd, J=9.3, 2.7Hz, 1H) ,
3.450 (br d, J=14.8 Hz, 1H) , 3.394 (s, 3H) , 3.332 (s,
20 3H) , 3.202 (s, 3H) , 2.709 (dd, J=17.4, 3.4Hz, 1H) ,
2.411 (dd, J=17.4, 8.4Hz, 1H) , 1.756 (s, 3H) , 1.657(s,
3H) , 1.061 <d, J=6.6 Hz, 3H) , 1.009 (d, J=6 . 6 Hz, 3H) ,
0.944 (d, J=6.6 Hz 3H) , 0.923 (d, J=6.6 Hz, 3H), 0.865
(d, J=6.7 Hz, 3H) , 0.648 (q, J=11.9 Hz, 1H) ; 13 C NMR
25 (CDCI3, 100.6 MHz, 3:1 mixture of trans :cis amide
rotamers/ data for the trans-rotamer): _ 211.9, 207.5,
196.1, 169.6, 166.2, 138.4, 138.1, 134.9, 132.3, 130.6,
127.3, 125.5, 125.2, 98.7, 84.4, 82.8, 76.2, 74.2, 73.9,
68.9; MS (ESl + /NH 4 OAc) m/z 936 (M+Na + ) , 931 (M+NH 4 + ) ; MS
30 (ESl"/NH 4 COOH) m/z 958 (M+HCOO") ; UV (MeOH) max 267,
277, 289 nm.
III. BIOLOGICAL EXAMPLES
35 Compounds of the invention were analyzed for
antifungal and immunosuppressive activity using the
following assays.
-35-
WO 94/02136
PCT/US93/06678
Assay for Antifungal Activity
Yeast organism (Saccharomyces cerevisiae) in
logarithmic growth were plated on complete agar medium
5 (YPD) . Compounds dissolved in an appropriate aqueous or
organic solvent were placed in wells punched in the
agar. Plates were incubated for 4 8 hours and zones of
inhibition were measured. All of the compounds of the
invention which were tested in this assay exhibited
10 antifungal activity.
Mitogenesis Assay for Immunosuppressive Activity
Spleen cells from BDF1 female mice were
established in RPMI with 10% fetal calf serum at 5 x
10^/mL. One hundred mL aliquots of this suspension (5 x
15 10^ cells) were dispensed into 96-well round-bottomed
microtiter plates (Linbro, Flow Laboratories) .
Concanavalin A (5 u g/ml)was added as the mitogenic
stimulus, and the final volume in the microtiter wells
was adjusted to 200 u L with RPMI. Cell cultures were
20 incubated for 72 hours at 37° C in a 5% CO2 atmosphere
and pulsed with 0.5 u Ci ^h— thymidine (specific activity
2.00 Ci/mole) for the last 18 hours of the 72 hour
culture. The cells were harvested on an automated
multiple sample harvester and cell-associated
25 radioactivity counted in a Beckman liquid scintillation
counter. The results are expressed as the mean values
derived from quadruplicate measurements. Cell viability
was determined by trypan blue exclusion after 72 hours
of incubation. Compounds to be tested were added to the
30 microtiter plates at the appropriate dilutions prior to
the addition of cells. All of the compounds of the
invention which were tested in this assay exhibited
immunosuppressive activity.
Results of these two assays, i.e., antifungal
35 activity assay and the mitogenesis assay for
immunosuppressive activity, for compounds of this
invention are provided in Table 1.
-36-
WO 94/02136
PCT/US93/06678
TABLE 1
Biological Activity
10
15
Compound of
Example #
l
2
* or V
•••iiiiii OEt
— OEt
Antifungal
l£i? ng/mL
<500
<500
Mitogenesis
_LLC^0__nMl
<500
<500
20
OM»
ovw <500
<500
25
4
5
6
• 0(CH 2 ) 2 OH <500
— -0(CH 2 ) 2 OH <500
— OAC <500
<500
<500
<500
<500
<500
30 8 H 2 <500 <500
9 SMe <500 <500
10 —SMe <500 <500
11 SPh <500 <500
13 allyl <500 <500
-37-
WO 94/02136
PCT/US93/06678
Table I - continued
Compound of
Example #
X or Y
Antifungal
Mitogenesis
— ac^n nM)
10
15
14
15
16
•wo"
3WO —
HO ■
HO 111111111
o=
<500
<500
<500
<500
<500
<1000
20
17
18
19
20
•wo-fc
<500
<500
<1000
<500
<500
<500
<500
<500
25
<500
While the above descriptions and Examples fully
30 describe the invention and the preferred embodiments
thereof, it is understood that the invention is not
limited to the particular disclosed embodiments coming
within the scope of the following claims.
-38-
WO 94/02136
PCT/US93/06678
CLAIMS
What is claimed is:
1. A compound of the formula
wherein :
X is selected from the group consisting of H,
-OR 10 , -S <0) n R 10 , -nrIOrII, alkyl and aryl;
Y is selected from the group consisting of H,
-OR 10 , -S(0) n R 10 , -NR 10 R 1:L , alkyl and aryl;
or X and Y taken together are =0;
n is selected from the group consisting of 0, 1
and 2;
R 1 is selected from the group consisting of =0,
(-OR 6 ,H) and (H,H);
R 2 is selected from the group consisting of =0,
(H, H) , and (H,OH);
R 3 and R 6 are independently selected from the
group consisting of -H, C1-C4 alkyl, -C(=0)R 7 ,
-C(=0)OR 7 , -C(=0)NHR 7 , and -C(=S)OR 7 /
R 4 is selected from the group consisting of =0
and (H,OR 6 );
or R 3 and R 4 can be taken together to form a
bridge of the formula A-C(R^) (R 9 ) -0-B, where A is a bond
to the oxygen bonded to the carbon at the 2 8-position
and B is a bond to the carbon at the 30-position;
r5 is selected from the group consisting of -H
-39-
WO 94/02136
PCI7US93/06678
and C1-C4 alkyl;
R 7 is selected from the group consisting of
c l"*ClO alkyl, C3-C5 cycloalkyl, aryl groups, and
heterocyclic groups;
5 and R^ are independently selected from the
group consisting of H, Cj to Cg alkyl, or R& and R^
taken together are =0/
R 1 ^ and R 11 are independently selected from the
group consisting of H, alkyl and aryl;
10 provided that,
(a) at least one of X and Y is H; and
(b) when Y is -R 10 , then R 10 is other than CH3;
and all pharmaceutically acceptable salts, hydrates or
solvates thereof.
15
2. A compound of Claim 1 where R 1 is (H,0H) .
3. A compound of Claim 1 where R^ is =0.
20 4. A compound of Claim 1 where R 3 is H.
5. A compound of Claim 1 where R 4 is =0.
6. A compound of Claim 1 where R 5 is H.
25
7. A compound of Claim 1 where R 1 is (H,0H), r2
is =0, R 3 is H, R 4 is =0, R 5 is H.
8. A compound of Claim 1 where one of X and Y is
30 selected from the group consisting of OR 1 ^ and SR 1 ^ or
where X and Y are taken together as =0.
9 . A compound of Claim 8 where X and Y are taken
together as =0.
35
10. A compound of Claim 8 where one of X and Y
is selected from the group consisting of -OR 10 and -SR 10
-40-
WO 94/02136
PCT/US93/06678
where R iU is selected from the group consisting of
optionally substituted C1-C3 alkyl, optionally
substituted benzyl, and optionally substituted phenyl.
11. A compound of Claim 1 where, when one of X
and Y is an alkyl group or when one of R 1 ^ and R 11 is an
alkyl group, said alkyl is selected from the group
consisting of branched, straight-chain, cyclic,
polycyclic, saturated and unsaturated alkyl groups which
may be substituted with 0, 1 or more substitutents
selected from the group consisting of aryl, keto,
hydroxyl, alkoxyl, acyloxy, amino, N-acylamino, halogen,
cyano and carboxyl substituents, and in which at least
one carbon atom may be replaced with a heteroatom
selected from the group consisting of O, S and N.
12. A compound of Claim 8 where, when one of X
and Y is an aryl group or when one of R 10 or R 11 is an
aryl group, said aryl group is selected from the group
consisting of cyclic, heterocyclic, polycyclic and
heteropolycyclic C2 to C14 aryl groups, which may be
substituted by one to five members selected from the
group consisting of alkyl, hydroxyl, alkyoxyl, acyloxy,
amino, N- acylamino, halogen, cyano, carboxyl and nitro
groups .
13. A compound of Claim 7 where one Of X and Y
is selected from the group consisting of OR^-0 and SR 1 ^
or where X and Y are taken together as =0.
14 . A compound of Claim 13 where X and Y are
taken together as =0.
15 . A compound of Claim 7 where one of X and Y
is selected from the group consisting of -OR 1 ^ and -SR 1 ^
where R^O is selected from the group consisting of
optionally substituted C1-C3 alkyl, optionally
WO 94/02136
PCI7US93/06678
substituted benzyl, and optionally substituted phenyl.
16. A compound of Claim 7 where, when one of X
and Y is an alkyl group or when one of R 10 and R 11 is an
5 alkyl group, said alkyl is selected from the group
consisting of branched, straight-chain, cyclic,
polycyclic, saturated and unsaturated alkyl groups which
may be substituted with 0, 1 or more substitutents
selected from the group consisting of aryl, keto,
10 hydroxyl, alkoxyl, acyloxy, amino, N-acylamino, halogen,
cyano and carboxyl substituents, and in which at least
one carbon atom may be replaced with a heteroatom
selected from the group consisting of O, S and N.
15 17. A compound of Claim 7 where, when one of X
and Y is an aryl group or when one of R 10 or R 11 is an
aryl group, said aryl group is selected from the group
consisting of cyclic, heterocyclic, polycyclic and
heteropolycyclic C2 to C14 aryl groups, which may be
20 substituted by one to five members selected from the
group consisting of alkyl, hydroxyl, alkyoxyl, acyloxy,
amino, N- acylamino, halogen, cyano, carboxyl and nitro
groups .
25 18. The compound of Claim 1 wherein:
(1) R 1 is (H,OH), R 2 is =0, R 3 is H, R 4 is =0,
R 5 is H, Y is H and X is -OCH3;
(2) R 1 is (H,OH), R 2 is =0, R 3 is H, R 4 is =0,
R 5 is H, Y is H and X is -0C 2 H 5 ;
30 (3) R 1 is (H,0H), R 2 is =0, R 3 is H, R 4 is =0,
R 5 is H, X is H and Y is -0C 2 H 5 ;
(4) R 1 is <H,0H), R 2 is =0, R 3 is H, R 4 is =0,
R 5 is H, Y is H and X is -OCH 2 CH 2 OH;
(5) R 1 is (H,0H), R 2 is =0, R 3 is H, R 4 is =0,
35 R 5 is H, X is H and Y is -OCH 2 CH 2 OH;
(6) R 1 is (H,0H), R 2 is =0, R 3 is H, R 4 is =0,
R 5 is H, Y is H and X is 3 , 4-dimethoxybenzyloxy ;
'-42-
WO 94/02136
PCI7US93/06678
(7) R 1 is (H, OH), R 2 is =0, R 3 is H, R 4 is =0,
r5 is H, X is H and Y is 3, 4-dimethoxybenzyloxy ;
(8) R 1 is (H,0H), R 2 is =0, R 3 is H, R 4 is=0, R 5
is H, Y is H and X is -SCH 3 ;
5 (9) R 1 is (H,OH), R 2 is =0, R 3 is H, R 4 is=0, R 5
is H, X is H and Y is -SCH 3 ;
(10) R 1 is (H,OH), R 2 is =0, R 3 is H, R 4 is=0,
R 5 is H, Y is H and X is -SPh where Ph is phenyl;
(11) R 1 is (H,OH), R 2 is =0, R 3 is H, R 4 is=0,
10 r5 is H, X is H and Y is -SPh where Ph is phenyl;
(12) R 1
is
(H,0H) , R 2 is =0,
R 3
u
n r
R 4
K
is
TJ
H,
Y is H
and
X is 2-furanyl;
(13) R 1
is
(H,OH) , R 2 is =0,
R 3
is
H,"
T>4
K
is=0,
r j
is
H,
X is H
and
Y is 2-furanyl;
15
(14) R 1
is
(H, OH) , R 2 is =0,
R 3
is
H,
0 4
R H
is=0,
R 5
is
H,
Y is H
and
X is O-acetyl;
(15) Rl
is
(H,0H) , R 2 is =0,
R 3
is
H,
R 4
is=0,
R5
is
H,
and each of X and Y is H;
(16) R 1
is
(H,OH) , R 2 is =0,
R 3
is
H,
R 4
is=0,
20
R5
is
H,
and X and
Y are taken together
as
=0
(17) Rl
is
(H, OH) , R 2 is =0,
R 3
is
H,
R 4
is=0,
R5
is
H,
X is H
and
Y is allyl;
(18) R 1
is
(H, OH) , R 2 is =0,
R 3
is
H,
R 4
is=0,
R5
is
H,
Y is H
and
X is OH;
25
(19) R 1
is
(H, OH) , R 2 is =0,
R 3
is
H,
R 4
is=0,
R5
is
H,
Y is H
and
X is 2, 4-dimethoxyphenyl;
(20) R 1
is
(H, OH) , R 2 is =0,
R 3
is
H/
R 4
is=0,
R5
is
H,
X is H
and
Y is 2, 4-dimethoxyphenyl;
or
(21) R 1
is
(H, OH) , R 2 is =0,
R 3
is
H,
R 4
is =C
30
R5
is
H,
X is H
and
Y is -OH.
-43-
WO 94/02136
PCT/US93/06678
10
19. A pharmaceutical composition comprising a
pharmaceutically acceptable carrier or diluent, and an
effective therapeutic or prophylatic amount of one or
more compounds of the formula
MeO'
R s O,
15 wherein :
X is selected from the group consisting of H,
-OR 10 , -S(0) n R 10 , -NR 10 R 1:L , alkyl and aryl;
Y is selected from the group consisting of H,
-OR 10 , -S(0) n R 10 , -NR 10 R 1:L , alkyl and aryl;
20 or X and Y taken together are =0;
n is selected from the group consisting of 0, 1
and 2;
R 1 is selected from the group consisting of =0,
(-OR 6 ,H) and (H,H);
25 R 2 is selected from the group consisting of =0,
(H,H), and (H, OH) ;
R3 and R^ are independently selected from the
group consisting of -H, - C4 alkyl, -C(=0)R 7 ,
-C(=0)OR 7 , -C(=0)NHR 7 , and -C(=S)OR 7 ;
30 R^ is selected from the group consisting of =0
and (H,OR 6 )/
or r3 and R 4 can be taken together to form a
bridge of the formula A-C(R 8 ) (R 9 )-0-B, where A is a bond
to the oxygen bonded to the carbon at the 28-position
35 and B is a bond to the carbon at the 30-position;
r5 is selected from the group consisting of -H
and C1-C4 alkyl;
-44-
WO 94/02136
PCT/US93/06678
R ' is selected from the group consisting of
C^-C^Q alkyl, C3-C5 cycloalkyl, aryl groups, and
heterocyclic groups;
and are independently selected from the
5 group consisting of H, to Cg alkyl, or R^ and R^
taken together are =0;
R 10 and R 11 are independently selected from the
group consisting of H, alkyl and aryl;
provided that,
10 (a) at least one of X and Y is H; and
(b) when Y is -OR 10 , then R 10 is other than CH 3 ;
and all pharmaceutically acceptable salts, hydrates or
solvates thereof.
15 20. The pharmaceutical composition of Claim 19
wherein :
(1) R 1 is (H,0H), R 2 is =0, R 3 is H, R 4 is =0,
R 5 is H, Y is H and X is -OCH3;
(2) R 1 is (H,0H), R 2 is =0, R 3 is H, R 4 is =0,
20 R 5 is H, Y is H and X is -0C 2 H 5 ;
(3) R 1 is <H,OH), R 2 is =0, R 3 is H, R 4 is =0,
R 5 is H, X is H and Y is -OC 2 H 5 ;
(4) R 1 is <H,OH), R 2 is =0, R 3 is H, R 4 is =0,
R 5 is H, Y is H and X is -OCH 2 CH 2 OH;
25 (5) R 1 is <H,OH), R 2 is =0, R 3 is H, R 4 is =0,
R 5 is H, X is H and Y is -OCH 2 CH 2 OH;
(6) R 1 is <H,OH), R 2 is =0, R 3 is H, R 4 is =0,
R5 is H, Y is H and X is 3, 4-dimethoxybenzyloxy;
(7) R 1 is (H,0H), R 2 is =0, R 3 is H, R 4 is =0,
30 r5 is H, X is H and Y is 3, 4-dimethoxybenzyloxy ;
(8) R 1 is <H,0H), R 2 is =0, R 3 is H, R 4 is=0, R 5
is H, Y is H and X is -SCH3;
(9) R 1 is <H,OH), R 2 is =0, R 3 is H, R 4 is=0, R 5
is H, X is H and Y is -SCH3;
35 (10) R 1 is (H,0H), R 2 is =0, R 3 is H, R 4 is=0,
R 5 is H, Y is H and X is -SPh where Ph is phenyl;
(11) R 1 is (H,0H), R 2 is =0, R 3 is H, R 4 is=0,
-45-
WO 94/02136
PCI7US93/06678
R 5 is H, X is H and Y is -SPh where Ph is phenyl;
(12) R 1 is (H,OH), R 2 is =0, R 3 is H, R 4 is=0,
R5 is H, Y is H and X is 2-furanyl;
(13) R 1 is (H,OH), R 2 is =0, R 3 is H, R 4 is=0,
5 R 5 is H, X is H and Y is 2-furanyl;
(14) R 1 is (H,OH), R 2 is =0, R 3 is H, R 4 is=0,
R 5 is H, Y is H and X is O-acetyl;
(15) R 1 is (H,OH), R 2 is =0, R 3 is H, R 4 is=0,
R5 is H, and each of X and Y is H;
10 (16) R 1 is (H,0H), R 2 is =0, R 3 is H, R 4 is=0,
R5 is H, and X and Y are taken together as =0;
(17) R 1 is (H,0H), R 2 is =0, R 3 is H, R 4 is=0,
R 5 is H, X is H and Y is allyl;
(18) R 1 is (H,0H), R 2 is =0, R 3 is H, R 4 is=0,
15 R 5 is H, Y is H and X is OH;
(19) R 1 is (H,0H), R 2 is =0, R 3 is H, R 4 is=0,
R5 is H, Y is H and X is 2, 4-dimethoxyphenyl;
(20) R 1 is (H,0H), R 2 is =0, R 3 is H, R 4 is=0,
R5 is H, X is H and Y is 2, 4-dimethoxyphenyl; or
20 (21) R 1 is (H,0H), R 2 is =0, R 3 is H, R 4 is =0,
R 5 is H, X is H and Y is -OH.
21 . A method of inhibiting the growth of
pathogenic fungi in a human or other animal in need
25 thereof which comprises administering to such human or
other animal an effective, non-toxic amount of a
composition of Claim 19.
22. A method of inducing immunosuppression in a
30 human or other animal in need thereof which comprises
administering to such human or other animal an
effective, non-toxic amount of a composition of Claim
19.
35 23. A method of treating carcinogenic tumors in
a human or other animal comprising administering to such
human or animal an effective, non-toxic amount of a
-46-
WO 94/02136
PCT/US93/06678
composition of Claim 19.
24. A method of preparing compounds of Claim 1
comprising contacting a compound of the formula:
5
10
where
15 • R 1 is selected from the group consisting of =0,
(-OR 6 ,H) and (H,H);
R 2 is selected from the group consisting of =0,
(H,H), and (H, OH) ;
R3 and R6 are independently selected from the
20 group consisting of -H, Ci - C 4 alkyl, -C(=0)R 7 , -
C(=0)OR 7 / -C(=0)NHR 7 , and -C(=S)OR 7 ;
R 4 is selected from the group consisting of =0
and (H,OR 6 );
or R 3 and R 4 can be taken together to form a
25 bridge of the formula A-C(R 8 ) (R 9 ) -O-B, where A is a bond
to the oxygen bonded to the carbon at the 2 8-position
and B is a bond to the carbon at the 30-position;
R5 is selected from the group consisting of -H
and Ci-C 4 alkyl;
30 R 7 is selected from the group consisting of C^-
Ciq alkyl, C3-C5 cycloalkyl, aryl groups, and
heterocyclic groups;
R 8 and R 9 are independently selected from the
group consisting of H, C'i to Cg alkyl, or R 8 and R 9
35 taken together are =0;
with an acid selected from the group consisting
of protic acids and Lewis acids, and with an appropriate
47-
WO 94/02136
PCT/US93/06678
nucleophile .
25. The method of Claim 24 where said acid is
selected from the group consisting of trif luoroacetic
acid and titanium tetrachloride.
26. The method of Claim 25 where said acid is
trif luoroacetic acid.
27. The method of Claim 25 where-R 1 is (H, OH) ,
r2 i s =0 , r3 i s H/ R 4 i s =0 , and R 5 is H.
28. The method of Claim 27 where said acid is
trif luoroacetic acid.
-48-
INTERNATIONAL SEARCH REPORT
International application No.
PCTAJS93/06678
A. CLASSIFICATION OF SUBJECT MATTER
IPC(5) :498/12; A61K 31/395
US CL :540/456 514/291
According to International Patent Classification (IPC) or to both national classification and IPC
B. FIELDS SEARCHED
Minimum documentation searched (classification system followed by classification symbols)
U.S. :
Documentation searched other than minimum documentation to the extent that such documents are included in the fields searched
NONE
Electronic data base consulted during the international search (name of data base and, where practicable, search terms used)
NONE
C. DOCUMENTS CONSIDERED TO BE RELEVANT
Category*
Citation of document, with indication, where appropriate, of the relevant passages
Relevant to claim No.
Y
Y
US, A, 5,118,677 (Caufield) 02 June 1992. See entire
document.
US, A, 5,102,876 (Caufield) 07 April 1992. See entire
document.
US, A, 5,023,263 (Von Burg) 11 June 1991. See entire
document.
1-8, 10-13, 15-
28
1-2,7,13,
15-28
1-8, 10-13, 15-
28
1, 3-6, 8,
10-12, 19,
21-22. 24
25-28
Further documents are listed in the continuation of Box C. | | See patent family annex.
•A'
"E*
"L"
■o-
■r
Special c a tcfocie e of cited i kwmiil K
ilnn—m dafinaia the srmiral etate of the art which i» not comidered
«o be part of particular relevance
»<l». <.n»ii.l|iiM 1 lil M/».>h.lli.iil Miilin..l fiUnt *****
document which may throw doubts on priority ckum(«) or which ■
cited to eetabliah the publication date of another citation or other
m (aa •pacified)
later document publiahed after the nlenatiaanl IUbic date or priority
date and not in conflict with the application but cited to inideritand the
principle or theory underiyint the mvcntaoa
document referring to an oral disclosure, uac exhibition or other
document published prior to the international filing date but later Ibaa
donncnt. of pellicular relevance: the claimed invention cannot be
considered novel or cannot be considered to involve an mvenuveeiep
when the docianent ie taken alone
document of particular relevance: the claimed invention cannot be
considered to involve an inventive atep when the document ie
t combined with t ™* other euch H«n— —w* ~^hi~ji~
being obvious to a peraon ekilled in the art
document B
mberof met
family
Date of the actual completion of the international search
02 SEPTEMBER 1993
Date of mailing of the international search report
OCT 14 1993 y ///
Name and mailing address of the ISA/US
Commiuioner of Patents and Trademark*
BoxPCT
Washington, D C. 20231
Facsimile No. NOT APPLICABLE
MARIANNE CINTINS f r L 1^ —
Telephone No. (703) 308-1235 "
Form PCT/ISA/210 (second sheet)(July 1992)*
INTERNATIONAL SEARCH REPORT
International application No.
PCT/US93/06678
Box I Observations where certain claims were found unsearchable (Continuation of item 1 of first sheet)
This international report has not been established in respect of certain claims under Article 17(2)(a) for the following reasons:
1. I I Claims Nog.:
because they relate to subject matter not required to be searched by this Authority, namely:
□
Claims Nos.:
because they relate to parts of the international application that do not comply with the prescribed requirements to such
an extent that no meaningful international search can be carried out, specifically:
3. Claims Nos.:
because they are dependent claims and are not drafted in accordance with the second and third sentences of Rule 6.4(a).
Box II Observations where unity of invention is lacking (Continuation of item 2 of first sheet)
This International Searching Authority found multiple inventions in this international application, as follows:
Please See Extra Sheet.
1 . I I As all required additional search fees were timely paid by the applicant, this international search report covers all searchable
claims.
2. | xj As all searchable claims could be searched without effort justifying an additional fee, this Authority did not invite payment
of any additional fee.
3 . || As only some of the required additional search fees were timely paid by the applicant, this international search report covers
only those claims for which fees were paid, specifically claims Nos.:
4. | | No required additional search fees were timely paid by the applicant. Consequently, this international search report is
restricted to the invention first mentioned in the claims; it is covered by claims Nos.:
Remark an Protest The additional search fees were accompanied by the applicant's protest.
| | No protest accompanied the payment of additional search fees.
Form PCT/ISA/210 (continuation of first sheet(l))(July 1992)*
INTERNATIONAL SEARCH REPORT
h»_niaUonal application No.
PCTAJS93/06678
BOX D. OBSERVATIONS WHERE UNITY OF INVENTION WAS LACKING
This ISA found multiple inventions as follows:
Group I, claims 1-20, 21 & 24-28, drawn to Rapamycin derivatives, their method of use in inhibition of the
growth of fungi, and a method for their synthesis, respectively, classified in Class 540, subclass 456 and Class 514,
subclass 183.
Group II, claim 22, drawn to a method of inducing immunosuppression utilizing a Rapamycin derivative,
classified in Class 514, subclass 183.
Group III, claim 23, drawn to a method of treating carcinogenic tumors utilizing a Rapamycin derivative,
classified in Class 514, subclass 183.
PCT Rule 13.1 states that the international application shall relate to one invention only or to a group of
inventions so linked as to form a single inventive concept (" requirement of unity of invention").
PCT Rule 13.2 states that unity of invention referred to in Rule 13.1 shall be fulfilled only when there is a
technical relationship among those inventions involving one or more of the same or corresponding special technical
features.
Annex B, Part 1(c), further defines independent and dependent claims. Unity of invention only is concerned
in relation to independent claims. Dependent claims are defined as a claim which contains all the features of another
claim and is the same category as the other claim. The category of a claim refers to the classification Of claims
according to subject matter, e.g. product, process, use, apparatus, means, etc.
Annex B, Part 1(e) indicates the permissible combinations of different categories of claims. Part l(e(i)) states
that inclusion of an independent claim for a given product, an independent claim for a process specially adapted for the
manufacture of the said product, and an independent claim for a use of the said product is permissible.
Herein, two additional independent claims, i.e. 22 & 23, are included in the application, and appear contrary
to PCT Rule 13.
Form PCT/ISA/210 (extra sheet)(July 1992)*
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